Dissertation Relationship of Gray Snow Mold

DISSERTATION

RELATIONSHIP OF GRAY SNOW MOLD DEVELOPMENT IN POA ANNUA/POA

PRATENSIS TO PERSISTENCE OF CHLOROTHALONIL AND FLUDIOXONIL

UNDER SNOW COVER AND EFFECT OF SNOW REMOVAL ON GRAY SNOW

MOLD DEVELOPMENT AT HIGH ALTITUDE GOLF COURSES.

Submitted by

Tamla Diann Blunt

Department of Bioagricultural Sciences and Pest Management

In partial fulfillment of the requirements

For the Degree of Doctor of Philosophy

Colorado State University

Fort Collins, Colorado

Fall 2012

Doctoral Committee:

Advisor: Ned Tisserat

Howard Schwartz Curtis Swift Tony Koski

ABSTRACT

RELATIONSHIP OF GRAY SNOW MOLD DEVELOPMENT IN POA ANNUA/POA

PRATENSIS TO PERSISTENCE OF CHLOROTHALONIL AND FLUDIOXONIL

UNDER SNOW COVER AND EFFECT OF SNOW REMOVAL ON GRAY SNOW

MOLD DEVELOPMENT AT HIGH ALTITUDE GOLF COURSES.

Gray snow mold caused by Typhula incarnata and T. ishikariensis (speckled snow mold) is a major problem on golf courses where snow cover persists for periods exceeding 60 days. The disease is primarily managed by preventive fungicide applications in fall prior to winter snow cover. However, fungicide rates necessary to control snow mold are often much higher than those needed to suppress other turf diseases during the summer. Studies were undertaken to determine the sensitivity of T. incarnata and T. ishikariensis isolates to chlorothalonil and fludioxonil, two fungicides commonly used for Typhula blight control. Growth of most isolates (70%) on potato dextrose agar (PDA) amended with 1 µg/ml chlorothalonil was inhibited by more than

50% relative to growth on non-amended PDA. However, almost all isolates exhibited at least some growth at concentrations as high as 500 µg/ml. Growth of most isolates was inhibited by more than 50% on PDA amended with 0.1 µg/ml fludioxonil and > 90% were inhibited by more than 80% at 1 µg/ml. The persistence of chlorothalonil and fludioxonil residues in the turf during winter was measured by gas chromatography/mass spectrometry. In 2005-2006, chlorothalonil concentrations decreased by approximately 50% the first week after application and then decreased at a rate of 0.7-1.0 µg/g tissue/day during snow cover. The rapid decrease in

ii chlorothalonil concentration after the first week was not observed in other years.

Instead, concentrations of chlorothalonil and fludioxonil in the verdure decreased at less than 1.0 µg/g tissue/day or remained nearly the same at most sampling dates indicating these fungicides did not dissipate rapidly under snow. Despite this, only marginal control of Typhula blight was observed in the fungicide-treated plots. In non-fungicide treated plots, Typhula blight patches developed rapidly between the December and

January sampling dates, sometimes to a level of plot damage equal to that observed in late April or May.

Golf course superintendents at high altitudes in Colorado apply fungicides in late

October before permanent snow cover to prevent gray or speckled snow mold development. Snow is also removed from putting greens in March and kept snow-free through spring to help suppress snow mold. However, the benefits of spring snow removal in snow mold suppression compared to potential turfgrass damage caused by exposure to low temperatures following removal have not been documented. We compared snow mold severity and turfgrass health on a Kentucky bluegrass fairway at

Breckenridge and annual bluegrass plots at Vail, CO with permanent snow cover through winter to plots where snow was removed from late October through mid-

November and to plots where snow was removed in mid-March and maintained snow- free through the spring. Both gray and speckled snow molds were observed at both locations and in each year, although the majority of the damage was caused by speckled snow mold. Snow removal influenced temperatures at the turfgrass surface during the winter and a similar trend was observed for each treatment in each year and at both locations.

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ACKNOWLEDGEMENTS

I would very much like to thank Dr. Ned Tisserat for his role as advisor and editor for my research project. His help, support and advice have been invaluable. I would also like to thank my committee, Drs. Howard Schwartz, Curt Swift and Tony Koski for their support and willingness to serve on my doctoral committee.

In addition, I would like to thank all the Tisserat lab members, past and present for their help with this project, as well as Galen Brunk from the Weed Science lab for his expertise on the GC/MS.

I thank all my friends and family who have taken this journey with me to its fruition. They all know who they are and how much their support means to me.

And finally, I would like to acknowledge the late Dr. Bill Brown, without whom this would never have happened. He encouraged me to continue my education and saw a potential in me that I didn’t see in myself at the time. His mentorship was without parallel.

TABLE OF CONTENTS

List of Tables ...... vi

List of figures ...... vii

CHAPTER 1 ...... 1 RELATIONSHIP OF GRAY SNOW MOLD DEVELOPMENT IN POA ANNUA/POA PRATENSIS TO PERSISTENCE OF CHLOROTHALONIL AND FLUDIOXONIL UNDER SNOW COVER ...... 1 Introduction: ...... 2 Methods and Materials: ...... 8 Results: ...... 16 Discussion ...... 23 LITERATURE CITED: ...... 47

Chapter 2 ...... 50 Introduction: ...... 51 Methods and Materials: ...... 53 Results ...... 57 Discussion ...... 61 LITERATURE CITED: ...... 88

APPENDIX 1 ...... 90 LIST OF SNOW MOLD ISOLATES ...... 90

APPENDIX 2 ...... 98 FUNGICIDE TRIAL RESULTS FOR BRECKENRIDGE, CO 2004-05 AND 2005-06 98 FUNGICIDE TRIAL RESULTS FOR VAIL, CO 2004-05, 2005-06, 2006-07, 2007-08, 2008-09, 2009-10, 2010-11...... 98

APPENDIX 3 ...... 117 OPTIMIZING THE GC/MS PROTOCOL ...... 117

LIST OF TABLES

Table 1.1. Typhula blight damage to a Kentucky Bluegrass fairway at Breckenridge, and an annual bluegrass fairway at Vail, CO, following fungicide applications...... 44 Table 1.2. Typhula blight damage to an annual bluegrass fairway at Vail, CO, in 2009 following applications of chlorothalonil and fludioxonil at labeled and 2X, 5X and 10X labeled rates...... 45 Table 1.3. Snow mold isolates identified by ITS sequencing. Biological species I is Typhula ishikariensis var. ishikariensis or T. ishikariensis var. idahoensis; Biological species II is T. ishikariensis var. canadensis...... 46

Table 2. 1 Comparison of one versus two fall fungicide applications for the control of Typhula blight at Vail and Breckenridge, CO 2005-2011...... 84

LIST OF FIGURES

Figure 1.1. Relationship between chlorothalonil residues in the verdure and sampling dates during the winter of 2005-06 following application of two rates of chlorothalonil to a Kentucky bluegrass fairway at Breckenridge, CO on 24 Oct 2006. The rates of chlorothalonil loss from the verdure from 14 Nov 2005 to 14 Mar 2006 were not different (P>0.10) between the high (-1.0 µg/g tissue/day, R2 = 0 .88, P<0.001) and low (-0.7 µg/g tissue/day, R2 = 0 .90, P<0.001) application rates of chlorothalonil. Shaded area represents period of snow cover during the winter...... 30 Figure 1.2. Relationship between chlorothalonil residues in the verdure and sampling dates during the winter of 2006-07 following application of two rates of chlorothalonil on 24 Oct 2006 to an annual bluegrass fairway at Vail, and a Kentucky bluegrass fairway at Breckenridge, CO. Rates of chlorothalonil loss from the verdure at Vail from 5 Nov 2006 2 to 26 Mar 2007 were not different (P >0.10) between the high (-0.3 µg/g tissue/day, R = 2 0.62, P =0.02) and low (-0.24 µg/g tissue/day, R =0.86, P =0.002) rates. Similarly chlorothalonil loss rates were not different (P>0.10) between high (-0.2 µg/g tissue/day, 2 2 R = 0.90, P = 0.001) and low rates (-0.09 µg/g tissue/day, R =0.76, P =.0 01) at Breckenridge. Shaded area represents period of snow cover during the winter...... 31 Figure 1.3. Relationship between chlorothalonil and fludioxonil residues in the verdure and sampling dates during the winter of 2006-2007 following application of the commercial fungicide mixture Instrata containing chlorothalonil (3.7 kg/ha), fludioxonil (0.006 kg/ha) and propiconazole (0.093 kg/ha), to an annual bluegrass fairway at Vail and a Kentucky bluegrass fairway at Breckenridge, Colorado. The rate of fludioxonil loss from the verdure 5 Nov 2006 to 26 Mar 2007 were not different (P >0.10) between 2 Vail (-0.004 µg/g tissue/day, R = 0.44, P = 0.09) and Breckenridge (-0.006 µg/g 2 tissue/day, R = 0.65, P = 0.03). Shaded area represents period of snow cover...... 32 Figure 1.4. Relationship between chlorothalonil and fludioxonil residues in the verdure and sampling dates during the winter 2007-08 following application of two rates of the commercial fungicide mixture Instrata containing chlorothalonil (3.0 and 3.7 kg/ha), fludioxonil (0.005 and 0.006 kg/ha) and propiconazole (0.076 and 0.093 kg/ha), and fludioxonil (0.008 kg/ha) applied alone (as the commercial fungicide Medallion 50WP) to an annual bluegrass fairway at Vail, Colorado. Shaded area represents period of permanent snow cover...... 33 Figure 1.5. Inhibition in diameter growth of Typhula ishikariensis isolates on potato dextrose agar amended with 1,10, 100, and 500 µg/ml chlorothalonil relative to growth in non-amended agar. A subset of isolates was screened at the higher concentrations...... 34 Figure 1.6. Inhibition in diameter growth of Typhula incarnata isolates on potato dextrose agar amended with 1, 10, 100, and 500 µg/ml chlorothalonil relative to growth in non-amended agar...... 35 Figure 1.7. Growth inhibition of Typhula ishikariensis on half-strength potato dextrose agar amended with 0.001, 0.01, 0.1 and 1 µg/ml fludioxonil relative to growth of isolates in non-fungicide amended agar...... 36

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Figure 1.8. Growth inhibition of Typhula incarnata isolates on half-strength potato dextrose agar amended with 0.001, 0.01, 0.1 and 1 µg/ml fludioxonil relative to growth of isolates in non-fungicide amended agar...... 37 Figure 1.9. Growth inhibition of Typhula ishikariensis isolates on half strength potato dextrose agar amended with 1 and 10 µg/ml propiconazole relative to growth in non- amended agar. A subset (57) of all isolates (179) was screened at the higher concentration...... 38 Figure 1.10. Growth inhibition of Typhula incarnata isolates on half strength potato dextrose agar amended with 1 and 10 µg/ml propiconazole relative to growth in non- amended agar...... 39 Figure 1.11. Concentration study (2008-09) using chlorothalonil applied at 10X the recommended rate showing that development of snow mold was inhibited; however, phytotoxicity damage to turfgrass was exhibited...... 40 Figure 1.12. Typhula blight damage to A) chlorothalonil treated (8.1 kg/ha), B) chlorothalonil treated (13.9 kg/ha), and C) an untreated plot on a Kentucky bluegrass fairway at Breckenridge, CO in May 2006...... 41 Figure 1.13. Typhula blight damage to A) chlorothalonil-treated plot (8.1 kg/ha), B) chlorothalonil treated plot (13.9 kg/ha), and C) an untreated plot on an annual bluegrass fairway at Vail, CO in March 2007...... 42 Figure 1. 14 Representative isolates of Typhula spp. Plate labeled 581 is Typhula incarnata; Plate labeled 411 is Typhula ishikariensis var. canadensis/BSII; Plate labeled 195 is Typhula idahoensis/BSI; and Plate labeled SM2 is Typhula ishikariensis var. ishikariensis/BSI. Note differences in color and size of the sclerotia...... 43

Figure 2.1. Golf Course superintendents at high elevation courses in Colorado routinely remove snow from putting greens in late February or early March...... 66 Figure 2.2. Annual bluegrass putting green after snow removal in late February 2006. Note that there is no discoloration or evidence of freeze injury to the turfgrass...... 67 Figure 2.3. Snow removal from plot in March 2008...... 68 Figure 2.4. Removal of soil cores with a 1.2-meter-long soil core remover during 2005- 06...... 69 Figure 2.5. Example of gray and speckled snow mold damage to turfgrass cores removed from snow-covered plots and incubated in greenhouse for 2 weeks (2006-07)...... 70 Figure 2.6. Effect of snow removal and fungicide applications on turfgrass damage at Vail and Breckenridge, CO 2005-06. The fungicide quintozene was applied 25 Oct prior to permanent snow cover on 21 Nov 2005. Snow was not removed (none), removed at weekly intervals during November (Fall), or removed at weekly intervals from 5 Mar, 2007 (Spring) until the final rating date on 7 May. Damage resulting from the November snow removal was a combination of winter injury and snow mold development. Means not followed by the same letter are significantly different (P =0.05) by Fisher’s LSD test...... 71 Figure 2.7. Effect of snow removal and fungicide applications on turfgrass damage at Vail and Breckenridge, CO 2006-07. The fungicide quintozene was applied 25 Oct prior to permanent snow cover on 7 Nov 2006. Snow was not removed at weekly intervals from plots (None), removed from 7 Nov to approximately the last week in November

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(Fall removal), or removed on 5 March, 2007 (Spring removal) until the final rating date on 7 May (presented here). Damage resulting from the November snow removal was a combination of winter injury and snow mold development. Means not followed by the same letter are significantly different (P =0.05)...... 72 Figure 2.8. Seasonal development of gray and speckled snow mold in non-fungicide treated plots on an annual bluegrass and Kentucky bluegrass fairway at Vail and Breckenridge, CO respectively in 2005-06. Shaded area represents the period of snow coverage on plots. Snow mold damage on all dates except 27 Apr 2006 were determined by removing turf cores from underneath the snow cover and estimating plot damage based on injury to the cores. Bars for each mean represent standard error. ... 73 Figure 2.9. Seasonal development of gray and speckled snow mold in non-fungicide treated plots on an annual bluegrass and Kentucky bluegrass fairway at Vail and Breckenridge, CO, respectively, in 2006-07. Shaded area represents the period of snow coverage on plots. Snow mold damage on all dates except 1 Apr were determined by removing turf cores from underneath the snow cover and estimating plot damage based on injury to the cores. Bars for each mean represent standard error. .. 74 Figure 2.10. Effect of snow removal and fungicide applications on turfgrass damage at Vail, CO 2007-08. The commercial fungicide Instrata (containing the active ingredients chlorothalonil, propiconazole and fludioxonil) was applied 25 Oct 2006 prior to permanent snow cover on 7 Nov. Snow was not removed from plots (None), or removed on 26 Mar 2008 (Spring removal) and weekly thereafter until the final rating date 24 Apr 2008. Means not followed by the same letter are significantly different (P =0.05) by Fisher’s LSD test...... 75 Figure 2.11. Temperatures at the turf surface on Kentucky bluegrass fairway at Breckenridge, CO during the winter 2005-06 for plots with no, fall (turquois), and spring snow removal (purple). Shaded area represents snow cover...... 76 Figure 2.12. Temperatures at the turf surface on annual bluegrass fairway at Vail, CO during the winter 2005-2006 for plots with no, fall (turquois), and spring snow removal (purple)...... 77 Figure 2.13. Temperatures at the turf surface on annual bluegrass fairway at Breckenridge, CO during the winter 2006-07 for plots with A) fall (brown line) and no snow removal (blue line) and B) spring (blue line) and no snow removal (brown line). The spring and fall snow removal bars indicate the period in which snow was physically removed from the plots at approximately weekly intervals...... 78 Figure 2.14. Snow mold damage was less severe on a Kentucky bluegrass tee at Breckenridge, CO where snow was compacted for Nordic track...... 79 Figure 2.15. Snow mold damage was less severe on a Kentucky bluegrass fairway at Breckenridge, CO where snow was compacted for Nordic track. Note turfgrass discoloration associated with snow compaction...... 80 Figure 2. 16 Temperatures at the surface of Kentucky bluegrass in adjacent areas of a fairway where snow had not been compacted or had been compacted for the construction of a Nordic ski track at Breckenridge CO, 2006-07...... 81 Figure 2.17. Temperatures at the surface of annual bluegrass in adjacent areas of a fairway where snow had not been compacted or had been compacted for the construction of a Nordic ski track at Vail CO, 2006-07...... 82

Figure 2.18. Damage to snow removal plots by snow mold at Vail, CO, 2007-2008. Plot 201 was no spring snow removal and no fungicide application, plot 203 was spring snow removal and no fungicide application. Plot 202 was no spring snow removal with a fungicide application and plot 204 was spring snow removal with a fungicide application...... 83

CHAPTER 1

RELATIONSHIP OF GRAY SNOW MOLD DEVELOPMENT IN POA ANNUA/POA PRATENSIS TO PERSISTENCE OF CHLOROTHALONIL AND FLUDIOXONIL UNDER SNOW COVER

SUMMARY

Typhula blight is a major problem on golf courses where snow cover persists for periods exceeding 60 days. The disease is primarily managed by preventive fungicide applications in fall prior to winter snow cover. However, fungicide rates necessary to control snow mold are often much higher than those needed to suppress other turf diseases during the summer. Studies were undertaken to determine the sensitivity of

Typhula incarnata and T. ishikariensis isolates to chlorothalonil and fludioxonil, two fungicides commonly used for Typhula blight control. Growth of most isolates (70%) on potato dextrose agar (PDA) amended with 1 µg/ml chlorothalonil was inhibited by more than 50% relative to growth on non-amended PDA. However, almost all isolates exhibited at least some growth at concentrations as high as 500 µg/ml. Growth of most isolates was inhibited by more than 50% on PDA amended with 0.1 µg/ml fludioxonil and > 90% were inhibited by more than 80% at 1 µg/ml. The persistence of chlorothalonil and fludioxonil residues in the turf during winter was measured by gas chromatography/mass spectrometry. In 2005-2006, chlorothalonil concentrations decreased by approximately 50% the first week after application and then decreased at a rate of 0.7-1.0 µg/g tissue/day during snow cover. The rapid decrease in chlorothalonil concentration after the first week was not observed in other years.

Instead, concentrations of chlorothalonil and fludioxonil in the verdure decreased at less than 1.0 µg/g tissue/day or remained nearly the same at most sampling dates indicating

1 these fungicides did not dissipate rapidly under snow. Despite this, only marginal control of Typhula blight was observed in the fungicide-treated plots. In non-fungicide treated plots, Typhula blight patches developed rapidly between the December and

January sampling dates, sometimes to a level of plot damage equal to that observed in late April or May.

Introduction:

Gray snow mold caused by Typhula incarnata and speckled snow mold caused by Typhula ishikariensis, collectively called Typhula blight when they occur together, are major problems on golf courses in areas where snow cover persists for long durations

(Hsiang et al., 1999). However, gray snow mold may also develop in a mild form during winter in the absence of snow cover (Smith, 1989). Symptoms of snow molds are evident at snowmelt and are usually exhibited as light yellow, straw-colored or grayish- brown turf from a few centimeters to a meter or more in diameter (Smith, 1989). Leaves in the affected areas are matted together and often are covered with grayish-white mycelium. The mycelium desiccates as the turf grass dries out leaving areas of what appears to be dead turfgrass. Most often, crowns of the turf plants are not affected and will produce new leaves within two-three weeks after snowmelt (Smiley, 2005). Gray snow mold will produce faintly pink sclerotia up to 5 mm in diameter and may be found on or within the infected tissues (Smith, 1989). They will darken with age to a reddish- brown or dark-brown, wrinkling as they dry. Sclerotia of gray snow mold can be variable in shape, ranging from subglobose to elongated, flattened or irregular (Smith, 1989).

Speckled snow mold develops where winters are longer and more severe, usually requiring a snow cover duration of at least 90 days and generally upwards of 120 days

(Chang et al., 2006). Sclerotia of speckled snow mold are readily detached from the leaf and are much smaller than gray snow mold. They can be subglobose or slightly flattened, light brown to almost black as they dry (Smith, 1989). A study done in

Wisconsin on the distribution of Typhula spp. found that in areas of longer snow cover days (northern Wisconsin), Typhula ishikariensis occurred more frequently and in areas of fewer days of snow cover (southern Wisconsin), T. incarnata was the dominant species of snow mold (Chang et al., 2006).

Typhula is a basidiomycetous genus which has been assigned to different taxonomic families over time (Hsiang et al., 1999). In the GenBank taxonomy web page

(http://www.ncbi.nlm.nih.gov/taxonomy), the genus Typhula is placed in the order

Thelephorales, family Typhulaceae (Hsiang et al., 1999). The classification of Typhula ishikariensis and its varieties have been through several iterations. Pathogenicity, distribution, basidiocarp size and mating experiments (Bruehl and Cunfer, 1975),

(Bruehl et al., 1975) split T. idahoensis from T. ishikariensis. Later studies regarded these two species as conspecific and designated T. idahoensis as a variety of T. ishikariensis and designated a variant of T. ishikariensis found in Canada as T. ishikariensis var. canadensis (Arsvoll & Smith, 1978). In 1980, Bruehl and Machtmes maintained that T. ishikariensis and T. idahoensis were separate species based on mating experiments between Typhula isolates. T. ishikariensis var. canadensis was rejected as a distinct variety (Bruehl and Machtmes, 1980). The T. ishikariensis complex

3 was divided into two biological species by Matsumoto; var. ishikariensis from turf in

North America belongs to biological species I (Matsumoto, 1982) and Biotype A (Hsiang et al., 1999), however, the Japanese biotype A (Biological Species I, Biotype A and

Group I) does not cause disease on turf under natural conditions; Biological species II

(Biotype C, Biotype Bss and Group II) includes the North American var. canadensis; and the Japanese biotype B, classified as Typhula idahoensis by Bruehl (Bruehl and

Cunfer, 1975), is also Group III and Biological species I (Hsiang et al., 1999). A publication by Hsiang using genetic markers found that, even though T. phacorrhiza, T. incarnata, and T. ishikariensis are distinct species, the varieties of T. ishikariensis do not show genetic differences at the species level (Hsiang & Wu, 2000). In 2006, the development of species specific PCR primers designed to identify T. incarnata, T. phacorrhiza, T. ishikariensis, T. ishikariensis var. ishikariensis and T. ishikariensis var. canadensis for samples collected from golf courses throughout Wisconsin were able to identify 1,835 isolates to one of the Typhula species and/or its variety (Chang et al.,

2006).

Cultural practices play an important role in the development of snow mold diseases. Grasses need nitrogen fertility in order to overwinter, but excessive nitrogen fertilization near the end of the growing season promotes lush growth and prevents the turfgrass from becoming winter hardy (Hsiang et al., 1999). However, fertilization with nitrogen after the growth of leaf blades has stopped in the fall may promote turfgrass recovery and rapid growth in the spring (Couch, 1995). Fertilization with inorganic fertilizers may reduce snow mold severity in Agrostis spp. as compared with using

4 organic fertilizers (Hsiang et al., 1999). Since gray and speckled snow mold requires prolonged snow cover, attempts have been made to reduce snow mold severity by removing snow from the turf surface (Hsiang et al., 1999) (see Chapter 2).

Recommendations for dealing with damage from snow molds in spring include improving drainage, replanting or re-sodding, promoting rapid drying and warming of the turf or lightly fertilizing the affected areas (Smiley, 2005).

The type of turfgrass also influences the severity of Typhula blight.

Bentgrasses, especially creeping bentgrasses (Agrostis palustris), are often severely damaged by Typhula blight (Smith, 1989, Vargas, 1994, Wang et al., 2005) found genotypic variation in creeping bentgrass clones to damage caused by T. ishikariensis.

Clones from fairways tended to be more resistant than those from putting greens.

Kentucky bluegrass (Poa pratensis) and annual bluegrass (Poa annua), often found on golf course fairways or putting greens in Colorado, are considered intermediate in susceptibility to Typhula blight (Vargas et al., 1972, Hsiang et al., 1999) although this conclusion appears to be based on observational, rather than experimental evidence.

Recent research into biocontrol and organisms antagonistic to Typhula spp. shows promise. Typhula phacorrhiza shows the most promise as a potential biocontrol.

Isolates have been identified that could inhibit disease caused by T. ishikariensis on creeping bentgrass. A culture filtrate of Pseudomonas flourescens was inhibitory to mycelial growth of T. ishikariensis and in field trials, a bacterial suspension was able to reduce snow mold damage (Hsiang et al., 1999). Trichoderma atroviride is a cold tolerant, versatile hyperparasite that was isolated from the sub-arctic region of Alaska

5 and has been found to suppress a wide range of economically important plant pathogens, including Typhula incarnata, T. idahoensis and T. ishikariensis (McBeath,

2002.). Introducing these antagonists, as well as establishment, has been the largest hurdle to practicing biocontrol. These antagonists have to be able to survive the harsh winter environment under the snow cover as well as be low-temperature tolerant.

Economics also factor into biocontrol applications. These treatments may need to be applied annually or bi-annually in order to build populations high enough to be antagonistic to snow mold pathogens. This may result in costs even greater than fungicide use, however environmental risks and societal concerns may justify these costs if efficacy of the biocontrol is acceptable (Hsiang et al., 1999).

Typhula blight on bentgrass or annual bluegrass putting greens and fairways in

Colorado is primarily controlled with fungicide applications applied just before snow cover in the fall. Nevertheless, Typhula blight can still produce varying degrees of damage to the golf course. In spring, after snow melt, Typhula blight damage can pose economic and aesthetic problems for the golf course superintendent to get the course in shape in time for spring play. During the 1970’s and early 1980’s, inorganic fungicides containing mercury and cadmium were widely and effectively used to control snow molds. However, due to their effect on people and their persistence in the environment, they fell into disfavor and their use was discontinued (Skoglund, 1998). The contact fungicide quintozene replaced the mercurial compounds as the product of choice for snow mold control because of its efficacy and price. However, quintozene is phytotoxic to creeping bentgrass (Vargas, 1994). In 2010 the use of quintozene was briefly halted

6 by the Environmental Protection Agency due to potential human health effects, but it was re-authorized for sale in August of 2011. Nevertheless, ongoing concerns about quintozene resulted in continued research into alternative fungicides for Typhula blight control.

Many other fungicide chemistries have been used in controlling Typhula blight

(Appendix 2). These include chlorothalonil, chloroneb, iprodione, fludioxonil, thiophanate methyl, vinclozolin, and some of the triazole and strobiluron fungicides

(Hsiang et al., 1999, Jung, 2007, Vargas, 1994,). In most cases a combination of active ingredients are used since several different fungal pathogens including Microdochium nivale, T. ishikariensis and T. incarnata may be active under snow cover (Hsiang et al.,

1999). The combination of chlorothalonil, fludioxonil and propiconazole has effectively controlled Typhula blight in Colorado (Appendix 3) and is widely used by superintendents. Chlorothalonil and fludioxonil are non-polar, broad-spectrum fungicides (Putnam et al., 2003), whereas propiconazole is a triazole. These fungicides are also sold in a commercially available formulation called Instrata (Syngenta

Professional Products, Greensboro, NC).

Fungicide rates for Typhula blight control usually are several times that required for controlling other turf diseases (Hsiang et al., 1999). The reason for this isn’t entirely understood, although factors such as insensitivity of fungal isolates to selected fungicides, chemical inactivity or effectiveness at low temperatures, fungicide dissipation from melting snow, or photo- or biodegradation during winter may play a role

(Monadjemi et al., 2011, Sigler et al., 2003, Stromqvist and Jarvis, 2005, Wu et al.,

2002). Fungicide dissipation or degradation is of particular interest, since a fungicide

7 applied in the fall must persist and remain at high enough concentrations, sometimes for periods of 4-5 months, to inhibit snow mold fungi. There have been many studies looking at the fate of fungicides after application to turf (Carroll, 1993, Carroll, 2001,

Sigler et al., 2003, Stromqvist and Jarvis, 2005), although these studies have been conducted during the summer when turfgrass is actively growing.

The objectives of this study were to identify the Typhula isolates collected from different golf courses in Colorado, determine the sensitivity of a subset of these isolates to chlorothalonil and fludioxonil, to monitor the persistence of these fungicides on the turf during the winter, and evaluate the efficacy of these products in controlling Typhula blight.

Methods and Materials:

Collection of Typhula isolates

Turf samples containing Typhula sclerotia were arbitrarily collected from snow mold patches that had developed in the fungicide-treated plots at Vail and Breckenridge during 2007 (Appendix 1). Individual sclerotia were disinfested in a 0.05% sodium hypochlorite solution for 30 seconds (Jo et al., 2006) then rinsed with sterile distilled water before plating on half-strength potato dextrose agar. Pure cultures were maintained on ½ PDA at 10°C. Typhula isolates were also collected from various locations in Colorado or were obtained from other sources (Appendix 1), and maintained on ½ PDA as described. Some isolates were collected from fairways that in past years had been treated with fungicides, including chlorothalonil, propiconazole and quintozene, whereas other isolates were collected from fairway roughs and other sites

8 where fungicides were less likely to have been applied. Typhula isolates were speciated by morphological characteristics of the sclerotia (Smith, 1989). The species of a smaller set of isolates (Table 3) was verified by amplifying the rDNA ITS region with the universal primers ITS4 and ITS5, sequencing the amplicon, then comparing it with known Typhula isolates using BLASTN (http://blast.ncbi.nlm.nih.gov/Blast.cgi). In all cases, the sequencing verified species identification based on morphology in culture.

Fungicide residue field plots

2005-06

To test the persistence of chlorothalonil in the turf through the winter, fungicide plots were established on a Kentucky bluegrass (Poa pratensis) fairway at the

Breckenridge Golf Course, Breckenridge, CO, on 24 Oct 2005. The grass was mowed at a height of approximately 1.5 cm during the growing season but was not mowed after fungicide application. Plots measuring 3 m by 3 m were left untreated or were sprayed with chlorothalonil (Daconil Ultrex 82.5WDG, Syngenta Professional Products,

Greensboro, NC) at 3.0 or 3.7 kg/ha or quintozene (Turfcide 400, American Vanguard

Corporation, Newport Beach, CA) at 15.12 kg/ha. Quintozene, a commonly used snow mold fungicide, was included for comparison of efficacy to chlorothalonil, but residues were not determined for this fungicide during the study. Applications were made using a CO2 sprayer pressurized to 138kPa and equipped with a spray boom with four 8004 flat fan nozzles to deliver water at 815 L per ha. Plots were arranged in a randomized complete block design with four replicates. The fairway was snow-free on the date of

9 application and fungicides were allowed to dry on the foliage before collecting the first samples.

Plot corners were marked with 2.4-m-long orange fiberglass poles such that the boundaries of each treatment could be identified during snow cover. A 6.35-cm- diameter X 5-cm-deep turf core sample was arbitrarily removed from each treatment using a 1.2-meter-long soil core remover. Samples were placed in plastic bags, labeled and stored at -80° C until they were processed for fungicide residue analysis. Samples were collected approximately weekly for the first month after application, biweekly for the following month, and monthly thereafter until the final collection on 2 May 2006.

2006-07

Fungicide trials were repeated in 2006 on a Kentucky bluegrass fairway at the

Breckenridge Golf Course and on an annual bluegrass fairway at the Vail Golf Club,

CO. The experimental design was identical to that in 2005-06, with an additional treatment of the commercial fungicide mixture Instrata 3.61SE (Syngenta Professional

Products, Greensboro, NC) containing 3.7 kg chlorothalonil, 0.006 kg fludiozonil and

0.093 kg propiconazole per ha. Fungicide applications were made 24 Oct 2006 at both locations. The turf had been mowed to a height of approximately 1.5 cm during the growing season but was not mowed following fungicide application. Sample removal and processing were the same as described above. Final sample removal and visual

10 evaluations at Vail Golf Club occurred on 26 March 2007, whereas final sample extraction and visual evaluations at Breckenridge was 1 May 2007.

2007-08

On 24 Oct 2007, plots on an annual bluegrass fairway at Vail were left untreated or treated with applications of fludioxonil (Medallion 50WP, Syngenta Professional

Products, Greensboro, NC) at 0.008 kg/ha, propiconazole (BannerMAXX 1.3ME,

Syngenta Professional products, Greensboro, NC) at 1.08 kg/ha or two rates of the

Instrata as used in 2006-2007. Sample collection and processing for fungicide residue analysis were the same as described above. Final samples were collected 12 May

2008.

Fungicide residues

Several extraction solvents and protocols were initially used to try to maximize extraction of chlorothalonil and fludioxonil from turf samples (Appendix 3). The final extraction procedure was based on research conducted by Mastovska (Mastovska and

Lehotay, 2004) using toluene as the solvent. Living, green turfgrass leaves and shoots, hereafter referred to as the verdure (Beard, 1973) were clipped at a height just above the thatch layer from the chlorothalonil or fludioxonil-treated turf cores collected at each sampling date. The thatch layer is defined as the tightly intermingled layer of dead and living stems and roots that develops between the verdure and soil surface (Beard,

1973). Three, 0.2 g (fresh weight) sub-samples of verdure from the core removed from each fungicide-treated plot were placed in 50 ml glass centrifuge tubes with Teflon-lined caps. Ten ml of water-saturated toluene and 10 ml of distilled water were added to each 50 ml tube. The tubes were agitated on a wrist-action shaker for 2 hours and then centrifuged for 20 minutes at 6000 x g and 4°C to separate the toluene from water. A

100 µl aliquot of the toluene phase containing the chlorothalonil extract was placed in a

2 ml volumetric tube to which 10 µl of a 0.1 mg/ml butylate internal standard solution and 1890 µl water-saturated toluene had been added. In the case of fludioxonil, 10 µl of

0.1 mg/ml metalochlor internal standard solution was added to 1990 µl of the toluene phase containing the fludioxonil extract (no dilution was needed). Tubes were then hand-shaken to mix the butylate and toluene or metalochlor and toluene; then approximately 1 ml of the mixture was transferred to vials for analysis by gas chromatography/mass spectrometry (GC/MS, Hewlett-Packard 5890 Series II Plus Gas

Chromatograph, Hewlett-Packard 5972 Series Mass Selection detector, Agilent

Technologies, Palo Alto, CA, USA).

Fungicide concentrations in the toluene phase were analyzed with the GC/MS by monitoring the masses for butylate (m/e 146,156,174) and chlorothalonil (m/e 264, 266,

268 270), or metolachlor (m/e 238, 240) and fludioxonil (m/e 248, 249). An HP 5MS 30 mm x 0.25 mm column was used with a flow of helium at 1.5 ml/min. The injection temperature for the chlorothalonil runs was 225°C and the detector temperature was

280°C. The program for detecting chlorothalonil was as follows: initial oven temperature 80°C (hold for 1 minute), ramped at 20°C min to 230°C, then held at 230°C

12 for 1.5 minutes with a run time of 16 min. For each sampling date, analytical samples of chlorothalonil at 50, 100, 250, 500 and 1000 ng/-ml-1 or fludioxonil at 0.25, 0.5, 1.0, 2.0 and 5.0 ng/-ml-1 in toluene were processed. The R2 values of the peak area ratio of chlorothalonil/butylate or fludioxonil/metalochlor to concentrations of chlorothalonil or fludioxonil, respectively, were always >97%.

Fludioxonil and chlorothalonil residues extracted from the 2007-08 Instrata treated plots were evaluated in a slightly different manner. Samples of verdure were prepared as above using the toluene/water in 50 ml glass tubes. A 100 µl aliquot of the toluene phase and 20 µl of a 0.5 mg/ml atrazine as the internal standard was placed in a 2 ml volumetric tube for the chlorothalonil extraction after which 1880 µl of toluene was added to bring to volume. A 1980 µl aliquot of the toluene phase was added to 20

µl of 0.5 mg/ml atrazine as the internal standard for the fludioxonil extraction from

Instrata. Fungicide concentrations in the toluene phase were analyzed with the GC/MS by monitoring the masses for atrazine (m/e 200, 215) and chlorothalonil (m/e 266) or fludioxonil (m/e 248, 249). An HP 5MS 30 mm x 0.25 mm column was used with a flow of helium at 1.5 ml/min. The injection temperature for the fungicide runs was 250°C and the detector temperature was 280°C. The program for detecting chlorothalonil was as follows: initial oven temperature 150°C (hold for 1 minute), ramped to 250°C at a rate of

5°/min with a total run time of 20 min. For each sampling date, analytical samples of chlorothalonil or fludioxonil at 0.05, 0.1, 0.25, 0.5, 1.0, and 2.0 µg/ml in toluene were processed. The R2 values of the peak area ratio of chlorothalonil/atrazine or

13 fludioxonil/atrazine to concentrations of chlorothalonil or fludioxonil, respectively, were always >99%.

Quality control (QC) samples consisted of verdure without fungicide or spiked with 50 or 100 µl of a 767 µg/ml chlorothalonil (made from the formulated Daconil Ultrex product) in 2005-06 and 2006-07. Formulated product was initially used because of the concern that inert ingredients might influence recovery. In 2006-07 and 2007-08 verdure spiked with a 0.5, 1 and 2 µg/ml of analytical fludioxonil or chlorothalonil were also included in each run. Mean recovery rates for chlorothalonil were 90.1% (n = 33,

SD ± 9.0) from Daconil Ultrex in 2005-06, 90.4 % (n=51, SD ± 9.6) from Daconil Ultrex in 2006-07, 90.2% (n=27, SD±7.9) from Instrata in 2006-2007 and 81.7% (n=24, SD ±

9.5) from Instrata in 2007-2008. Mean recovery rates for fludioxonil were 94.2% (n=9,

SD±3.4%) from Instrata in 2006-07 and 94.1% (n=32, SD ± 5.9) from Instrata and

Medallion in 2007-08.

Data from all GC/MS were converted to fungicide concentrations (µg/g) using the ratio of butylate to chlorothalonil, metolachlor to fludioxonil or atrazine to chlorothalonil/fludioxonil and then to µg/g fresh plant tissue. Linear regression analysis

(SAS Inc., NC) was performed between fungicide concentrations and sampling dates.

To more accurately assess fungicide dissipation under snow cover, the initial sampling date as well as those that occurred more than two weeks after snow melt in the spring, were omitted from the final regression analysis.

Efficacy of chlorothalonil and fludioxonil at high field application rates

Field studies from 2005-06, 2006-07 and 2007-08 indicated that labeled rates of chlorothalonil and fludioxonil applied to the fungicide plots did not control Typhula blight

(Figure 1.12 and 1.13, Table 1.1). Therefore, the possibility of controlling Typhula blight using higher fungicide rates (i.e. rates 2, 5 and 10 times the concentration of the highest labeled rate) was evaluated in an annual bluegrass fairway at Vail in 2008-09. The fairway had been maintained at a height of 1.5 cm prior to the experiment, but was not mowed following the fungicide applications. Chlorothalonil (Daconil Ultrex) at 12.5,

22.5, 63.0 and 126.0 and fludioxonil (Medallion) at 0.38, 0.77, 1.91 and 3.86 kg/ha were applied on 29 Oct 2008 in a randomized complete block design with four replications.

Applications were made in the manner previously described. Plots were rated 27 Apr

2009 for Typhula blight severity. Residue samples were not collected during the winter and the experiment was not repeated.

In vitro fungicide inhibition assays

A 7 mm plug of agar containing mycelium of one of 180 Typhula ishikariensis or

15 T. incarnata isolates was placed individually in the center of a petri plate containing

½ PDA amended with chlorothalonil (Daconil Ultrex) at 0 and 1 µg/ml. An arbitrarily selected subset of T. ishikariensis (57 isolates at 10 and 100 µg/ml and 20 isolates at

500 µg/ml) and T. incarnata (11 isolates at 10, 100 and 500 µg/ml) was tested at other concentrations. These concentrations were selected because they were in the range of chlorothalonil residues detected in the verdure of fungicide-treated plots. Growth T.

15 ishikariensis was determined in ½ PDA plates amended with fludioxonil (Medallion) at 0,

0.1 and 1.0 µg/ml (199 isolates) or 0.01 and 0.001 µg/ml (39 isolates). Fifteen T. incarnata isolates were tested at the same concentrations. Growth was recorded of 179

T. ishikariensis and 15 T. incarnata isolates in agar amended with propiconazole

(BannerMAXX) at 0 and 1 µg/ml. A subset of 57 T. ishikariensis and 11 T. incarnata isolates were also grown on agar amended with propiconazole at 10 and 100 µg/ml. All plates were incubated in a growth chamber with no internal light at 10°C for 22 or 30 days, and radial growth of the fungus on the plates was measured. The radius of the initial agar plug was subtracted from each measurement. The percent relative mycelial growth (RMG) at each fungicide concentration was calculated as the radial growth (on fungicide-amended ½ PDA/radial growth or non-amended ½ PDA) x 100 (Jo et al.,

2006). The relationships between RMG and fungicide concentrations were analyzed by linear regression analysis (Minitab 14). However, R2 values were often low (< 0.80) and there was no linear relationship at the fungicide concentrations we tested for many isolates. Therefore EC50 values were not determined by regression analysis. Instead the percentage of growth inhibition of each isolate at each fungicide concentration was determined.

Results:

Typhula isolates

A systematic survey of the distribution and frequency of Typhula species in

Colorado golf courses was not conducted, but isolates were collected from several

16 locations in the state (Appendix 1). T. ishikariensis was present at most locations whereas T. incarnata and T. phacorrhiza were less frequently recovered. A more intensive collection was made from within plots at Vail and Breckenridge and 467 of the isolates collected were identified as T. ishikariensis based on morphological characteristics of their sclerotia. Although T. ishikariensis was dominant at both sites, eleven isolates arbitrarily collected from an annual bluegrass rough adjacent to the fungicide plots at Vail in 2007 were identified as T. incarnata.

A subset of 110 Typhula isolates were further characterized based on rDNA ITS sequencing results (Table 1.3). Of these, 52% of the isolates were identified as T. ishikariensis biological species 1 and 19% were identified as T. ishikariensis biological species II.

In addition to Typhula species, other fungi associated with snow mold damage were collected. Myriosclerotinia borealis was identified at a single site in Wolcott, CO,

Waitea circinata was recovered from damaged turf at Granby, CO and Fibularhizoctonia carotae was associated with mold on leaf debris on fairways at Loveland and Fort

Collins, CO. Another fungus that was apparently associated with snow mold damage on Kentucky bluegrass fairways and that developed sclerotia resembling Typhula was collected from Loveland and Colorado Springs. However, the rDNA ITS sequence was less than 80% similar to any of the Typhula species and it remained unidentified.

Residue trials

2005-06

Chlorothalonil residues in the verdure immediately following application of the low and high rates of Daconil Ultrex on 24 Oct 2005 were 132 and 75 µg/g tissue, respectively (Figure 1.1). Residues decreased by >50% after the first week and during a period of no snow cover. Snow covered the plots after the first week in November and remained for 134 consecutive days. Chlorothalonil residues decreased at a much slower rate of -0.7 (R2 = .90, P <0.001) and -1.0 (R2 = .88, P <0.001) µg/g tissue/day in the low and high Daconil Ultrex-treated plots, respectively, during this period. On 14

Mar 2006, the sampling date just prior to spring snow melt, chlorothalonil concentrations of 7 and 24 µg/g tissue at the low and high application rates were detected in the verdure. Chlorothalonil residues also were detected at levels <20µg/g tissue in samples collected after snowmelt. Despite continued presence of chlorothalonil residues in the verdure throughout the winter, chlorothalonil-treated plots sustained >50% damage from

Typhula spp. (Table 1.1).

2006-07

Chlorothalonil residues in the verdure immediately following applications of

Daconil Ultrex at the low and high rates on 24 Oct 2006 were 54 and 97, and 27 and 66

µg/g tissue at Vail and Breckenridge, respectively (Figure 1.2). Residues at

Breckenridge were similar to those detected during the initial sampling date at the same site in 2005 (Figure 1.1). Unlike 2005, there was no rapid decrease in chlorothalonil

18 residues the first week after application at either site. Instead, slightly higher chlorothalonil concentrations were measured after the first week at Vail. There was no permanent snow cover at either location until mid-November 2006 and approximately the top 1.5 cm of the soil froze during this period. The soil subsequently thawed after permanent snow cover as determined by verdure temperatures recorded by thermistors during the winter (data not shown). Snow melt occurred during the first week in Mar

2007 after which light rain and snow continued to fall through May, but any snow that fell was not permanent on the plots. There was consistently higher chlorothalonil residues on the annual bluegrass plots at Vail compared to the Kentucky bluegrass plots at Breckenridge throughout the permanent snow cover period. Chlorothalonil

2 concentrations from 1 Nov 2006 through 26 March 2007 decreased at a rate of -0.2 (R

= .86, P=0.002) and -0.3 (R2=.62, P=0.02) µg/g tissue/day at Vail and -0.1 (R2 = .76,

2 P=0.01) and -0.2 (R = .90, P = 0.001) at the low and high application rates, respectively, (Figure 1. 2), and chlorothalonil concentrations at the end of this period were 11 and 24, and 25 and 59 µg/g tissue for the low and high chlorothalonil application rates at Breckenridge and Vail, respectively. Similar chlorothalonil residues at both sites were measured on 12 May 07. As in 2005-06, all plots at both locations treated with the two rates of chlorothalonil sustained injury from Typhula spp. (Table

1.1), although damage was much more severe on the annual bluegrass fairway at Vail

(50-80%) compared to Breckenridge (24-40%).

In 2006, chlorothalonil was also applied to plots as part of the formulation of the commercial fungicide mixture Instrata. The chlorothalonil application rate (3.7 kg/ha) in the mixture was approximately half that of the low application rate of Daconil Ultrex (8

19 kg/ha) in the same trial. Nevertheless, chlorothalonil concentrations in the verdure at most sampling dates were similar (Figure. 1.3). Chlorothalonil concentrations in the

Instrata-treated plots were consistently lower at Breckenridge than Vail. Chlorothalonil concentrations in the verdure of Instrata-treated plots were consistently lower at

Breckenridge than Vail. There was no clear trend in chlorothalonil residue changes in the verdure during snow cover at either site (Figure 1.3). For example, chlorothalonil concentrations from the Instrata application were 31 and 44 µg/g tissue on 24 Oct 2006 compared to 28 and 61 µg/g tissue on 26 Mar 2007 at Breckenridge and Vail, respectively. Residues of fludioxonil, another component of the Instrata fungicide, were

<4µg/g tissue immediately following applications at both sites. There was a slight trend toward decreasing concentrations through the winter, although fludioxonil could be detected at all sampling dates. Plots treated with Instrata had approximately 8%

Typhula blight damage at Breckenridge compared to 48% at Vail (Table 1.1).

2007-08

On 24 Oct 2007, chlorothalonil was applied to plots at Vail as part of the commercial fungicide mixture Instrata. Chlorothalonil residues in the verdure immediately after application were 57 and 75 µg/g tissue for the low and high Instrata rates, respectively, and did not change appreciably after the first week (Figure 1.4).

There was no permanent snow cover until 20 Nov 2007, and the verdure and top 1.5 cm of soil froze by 15 Nov. The soil and verdure thawed following permanent snow cover.

Snow melt occurred during the first week in May 2008. Chlorothalonil residues

20 fluctuated during the winter with no clear trend until 1 Apr 2008 when concentrations at the two application rates dropped to approximately 25 µg/g tissue. Initial fludioxonil residues in verdure resulting from low and high Instrata application rates were 0.5 and

0.8 µg/g tissue, respectively, and residues below 1.1 µg/g tissue were detected Until 1

Apr (Figure 1.4). No fludioxonil residue was detected in May at either application rate.

Application of fludioxonil alone (Medallion) to plots at 1.0 kg/ha resulted in much higher initial residues (13 µg/g tissue) than Instrata applications. Residues remained above 10

µg/g tissue until 6 Mar and then decreased rapidly at the last two sampling dates. Plots treated with fludioxonil only (Medallion 50WP) sustained more (76%) Typhula blight damage than plots treated with a combination of fludioxonil, chlorothalonil and propiconazole (Instrata) at the low (36%) and high (22%) application rate (Table 1.1).

Efficacy of chlorothalonil and fludioxonil at high application rates

Because labeled rates of chlorothalonil and fludioxonil failed to provide adequate

Typhula blight control in the residue experiments, applications of up to 10X the highest labeled rates of these fungicides were applied. Typhula blight was not effectively controlled at any rate of fludioxonil (Table 1.2). Only the highest rate of chlorothalonil provided some measure of control (14% damage), but this rate also caused phytotoxicity to the annual bluegrass (Figure 1.11).

In-vitro fungicide inhibition assays

Chlorothalonil was inhibitory to the growth of T. ishikariensis to some extent at all concentrations tested (Figure 1.5). Although growth in 21% of the isolates was inhibited by >90% at 1 µg/ml chlorothalonil, 53% (96/180) had a growth reduction <80%.

Increasing the chlorothalonil concentrations to 10 and 100 µg/ml increased growth inhibition in some of the isolates, but not in others. For example, the proportion of isolates that were inhibited by >90% remained similar at 10 (22%), 100 (19%) and 500

(18%) µg/ml. Thus increasing the chlorothalonil concentration to 500 µg/ml did not result in complete inhibition of all isolates. Typhula incarnata isolates were even less sensitive to chlorothalonil. Twelve of 15 isolates were only slightly or not inhibited

(<10%) at 1 µg/ml, and only 3 of 11 isolates had a growth inhibition >90% at a concentration of 500 µg/ml (Figure 1.6).

Growth in approximately half of the T. ishikariensis isolates was slightly or not inhibited (<10%) at 0.001 and 0.01 µg/ml fludioxonil, and none were completely inhibited at these concentrations (Figure 1.7). Increasing the fludioxonil concentration to 1 µg/ml resulted in growth reductions >80% in a larger proportion (85%) of isolates.

Nevertheless, only 12% of the isolates were completely inhibited or nearly so (>90%) at

1 µg/ml. Growth reduction in T. incarnata followed a similar pattern with very little inhibition (<10%) in all isolates at 0.001 µg/ml, increasing inhibition at 0.01 and 0.1

µg/ml, but still not complete inhibition at 1 µg/ml (Figure 1.8). Growth of all T. ishikariensis and T. incarnata isolates were completely inhibited at 10 µg/ml fludioxonil

(data not shown). Propiconazole at 1µg/ml reduced growth by varying amounts in T.

22 ishikariensis isolates, but only 3 isolates were inhibited >90% (Figure 1.9). Although growth was further reduced in many isolates at 10 µg/ml, only 3 additional isolates were inhibited by >90%. Growth inhibition in T. incarnata followed a similar pattern with increasing but not complete inhibition at 10 µg/ml propiconazole (Figure 1.10). Growth of all T. ishikariensis and T. incarnata isolates were completely inhibited at 100 µg/ml propiconazole (data not shown).

Discussion

The collection of Typhula ishikariensis and T. incarnata from many sites in

Colorado is in agreement with previous studies (Chang et al., 2006, Smith, 1989) that showed these species are widely distributed in North America. There was a general trend for T. ishikariensis to be more widely distributed at high elevations in Colorado

(Vail, Breckenridge), although this may have been an artifact of the sampling procedure.

Also, both species were recovered from golf courses at lower elevations (Ft. Collins and

Loveland).

Only T. ishikariensis was recovered from the fungicide plots at Vail and

Breckenridge in a more intensive sampling in 2007, even though T. incarnata was consistently collected from blighted turfgrass in the fairway rough less than 100 m away.

The dominance of T. ishikariensis on bentgrass greens and fairways has also been observed in Wisconsin and Utah (Chang et al., 2006). The reason for this isn’t known, but it is unlikely the apparent exclusion of T. incarnata from the fungicide plots was a result of environmental differences such as snow depth or length of snow cover. It is also unlikely that a difference in fungicide sensitivity between the species is the cause,

23 even though the roughs are rarely treated with fungicides and fairways are treated annually. Typhula incarnata was similar in sensitivity to propiconazole, and less sensitive to fludioxonil and chlorothalonil than T. ishikariensis. T. incarnata is considered a weak pathogen, but is better able to survive in a saprophytic phase than T. ishikariensis (Matsumoto, 1994). It also adapts to less favorable locations for snow mold development (Smith, 1992) with a major saprotrophic phase. In contrast, the more aggressive T. ishikariensis may be better able to colonize highly maintained turfgrass on the fairway and thus displace T. incarnata from these sites.

During the general survey, a Typhula-like fungus was recovered from blighted

Kentucky bluegrass patches in Colorado Springs, Fort Collins and Loveland. The fungus produced sclerotia with somewhat similar characteristics to T. incarnata. The rDNA ITS sequences of the isolates from the different locations were the same, but were distinctly different from published sequences of T. incarnata and T. ishikariensis.

However, the sequences matched (>98% similarity) those of an unidentified snow mold fungus collected in Oklahoma (Conway and Williams, 1986) and Wisconsin (Millett,

1999). The fungus was slightly pathogenic to creeping bentgrass in a Wisconsin study

(Millett, 1999), whereas no evidence of pathogenicity was detected in laboratory studies conducted in Oklahoma (Conway and Williams, 1986) and Colorado (Jorge Ibarra and

Ned Tisserat, Colorado State University, personal communication). The fungus probably has a mostly saprophytic phase colonizing dead turfgrass damaged by

Typhula blight

This general survey also correlated with the survey that Chang et al. (Chang et al., 2006) conducted in Wisconsin, Minnesota and Utah to identify Typhula species from

24 golf courses in these states. They found that when there were an increased number of snow cover days, the frequency of Typhula ishikariensis increased. This is very similar to what was found in the survey of golf courses in Colorado. Golf courses in the mountains of Colorado generally have upwards of 120 days of snow cover each winter, and the primary species that was recovered from these areas was Typhula ishikariensis

(Biological species I). The frequency of Typhula ishikariensis may also be due to its’ greater inoculum potential (Chang et al., 2006), as well as environmental conditions.

Fungicides have not always provided the desired level of control of Typhula blight even though they may be applied at very high rates ( Appendix 2), (Hsiang et al., 1999).

One possible explanation is that fungicides may dissipate in the verdure prior to snow cover or during the long period of snow cover. Wu et al. (2002) found that chlorothalonil persisted in the turf for only 12 days, with a dissipation half-life of 4.9 days, following application to a bentgrass putting green in summer. Mondadjemi et al.

(2011) reported a similar simulated half-life of 5.3 days for chlorothalonil and concluded photodegradation played an important role. Thus, applying a fungicide too far in advance of snow cover might result in reduced fungicide residues in the verdure.

However, I did not observe significant (>50%) decreases in chlorothalonil residues, except at Vail in 2005, following application; even though, in some cases, fungicides were applied up to three weeks before snow cover. Furthermore, chlorothalonil and fludioxonil persisted throughout the winter under snow cover with minimal loss in concentration.

The lack or slow rate of chlorothalonil and fludioxonil dissipation during late fall and early winter may be a result of several factors. Fairways were not mowed, and as a

25 result, clippings containing fungicide residue were not dislodged to the thatch or soil following fungicide application. Both bio- and photodegradation can be a factor in fungicide dissipation (Monadjemi et al., 2011, Sigler et al., 2003, Wang et al., 2011 ), but cold temperatures inhibiting microbial growth, shorter day lengths, and a diminished light intensity under snow likely reduced losses by these factors. Fungicides can also be washed from the foliage. Carroll et al. (Carroll, 1993 and 2001) found that chlorothalonil washoff was relatively slow and constant once the fungicide dried; however, significant loss could occur during the first hour if a simulated rainfall occurred. Any precipitation at high elevations in Colorado following fungicide application would likely occur as snow.

Melting snow should not have the same washoff potential as rainfall. It was observed that the ground beneath the snow remained frozen during winter and a thin film of water was present on leaves. It was also noted on the samples that were collected that the turfgrass continued to grow, as evidenced by the new shoots of annual bluegrass or

Kentucky bluegrass. This apparently was not sufficient to dislodge or reduce a significant amount of fungicide during the winter. A practical implication of these results is that while it is still desirable to apply fungicides as close to snow cover as possible, applications within 1-2 weeks before permanent snow should not cause significant fungicide dissipation.

Chlorothalonil and fludioxonil concentrations in the verdure immediately following application in each year were approximately 8-12 times lower than predicted (880 and

90 µg/g tissue for the highest chlorothalonil and fludioxonil rates, respectively) based on calculations using the average weight of the verdure collected from the turf cores. Wu et al. (Wu et al., 2002) also observed an eight fold difference between observed and

26 predicted concentrations of chlorothalonil on a bentgrass green. In their study, approximately 200 µg chlorothalonil/g tissue were recovered immediately after application of the fungicide at a rate of 24.5 kg/ha compared to the 68-115 µg/g tissue I recovered following application of 13.9 kg/ha (44% less) of the fungicide. In contrast,

Carroll et al. (2001) reported recovery of about 5,000 µg/g chlorothalonil in foliage following application of the fungicide at 9.2 kg/ha.

The reason for the apparent discrepancies between theoretical and observed fungicide residues isn’t known. Chlorothalonil is a lipophilic compound and presumably penetrates waxy leaf surfaces making extraction more difficult. Nevertheless, extraction efficiency from bluegrass leaves spiked with known concentrations of chlorothalonil was always greater than 81% in my studies. The turfgrass is not a flat surface, and it is possible some fungicide did not adhere to leaves during application. For example,

Carroll et al. (Carroll, 2001) estimated that only 51-55% of chlorothalonil was intercepted in the turfgrass canopy. However Wu et al. (Wu et al., 2002) found only a very small concentration of chlorothalonil was present in thatch one day after application, and no detectable amounts were found in soil. In the preliminary GC optimization runs, there was no detection of chlorothalonil in thatch samples, although the sampling was not extensive.

All of the fungicides tested were inhibitory to T. ishikariensis and T. incarnata in vitro. However, while concentrations of chlorothalonil as low as 1 µg/ml inhibited most of the isolates to some degree, a majority of isolates exhibited at least some growth at concentrations as high as 500 µg/ml. Similarly, most T ishikariensis and T. incarnata isolates exhibited at least some in vitro growth at fludioxonil and propiconazole

27 concentrations detected in the field. There was no evidence of a ‘qualitative’ resistance whereby an isolate exhibited complete insensitivity to the fungicide (Deising et al., 2008)

. Instead, isolates exhibited more ‘quantitative’ effects in which they may tolerate increasing concentrations of fungicide until some critical threshold is reached. Efflux transporters in fungal membranes are responsible for removing toxic chemicals (e.g. phytoalexins) from their cells (Deising et al., 2008) and it is possible that this mechanism is reducing fungicide levels in the fungal cells to sub-lethal levels.

I conducted the in vitro assays at 10°C and it is possible that results would have been different had they been done at 1°C. Typhula species are psychrophilic and are more metabolically active at low temperatures than most fungi. The possible combination of slower absorption of the fungicide and an active detoxification or efflux system might have allowed the fungus to tolerate relatively high fungicide concentrations.

The fungicide concentrations used in the in vitro studies were similar to those observed in the turfgrass verdure in field plots. Given that snow cover may persist for 4 months or longer, these data suggest that even limited fungal growth could result in significant turf colonization. For example, T. ishikariensis isolates had an approximate diameter growth of 3 mm/day on non-amended agar at 10°C. Even with a 50% growth reduction resulting from fungicide inhibition, a Typhula colony would attain a diameter of

18 cm after 4 months. Obviously many other factors, including near freezing temperatures under the snow could influence the growth rate, but it is plausible that a growth-inhibited fungus could cause damage to the turfgrass provided the fungicide did not affect pathogenicity. A high sclerotial population in the fungicide-treated turfgrass,

28 and subsequently the development of many small infection centers, could still result in major damage unless fungicide concentrations in the verdure exceeded that needed for complete inhibition. This may explain why increasing fludioxonail and chlorothalonil application rates in the field did not result in complete control of Typhula blight.

But even at the higher concentrations of fludioxonil and chlorothalonil, Typhula blight was not totally inhibited. And in the in-vitro studies, with higher concentrations of fludioxonil, chlorothalonil and propiconazole, it was noted that what appeared to be sporocarps were growing on the original mycelial plug, although this was never confirmed. It appeared that the fungus was trying to grow away from the fungicide- amended agar. So it may be that the fungus is able to ‘grow out’ of the fungicide layer in the turfgrass in order to cause disease. These structures were not observed in the field or on any of the collected samples, but they may not have survived for very long in the verdure. Since both fludioxonil and chlorothalonil are contact fungicides, new turf growth under snow cover may also reduce fungicide efficacy. Even though recovery of sufficient amounts of fungicide residue from the verdure should theoretically inhibit fungal growth, this was not inhibitory to the fungus. This may be why, in the funigicide trials (Appendix 2), two or more fungicides combined in one application seem to work better than individual compounds.

Figure 1.2. Relationship between chlorothalonil residues in the verdure and sampling dates during the winter of 2006-07 following application of two rates of chlorothalonil on 24 Oct 2006 to an annual bluegrass fairway at Vail, and a Kentucky bluegrass fairway at Breckenridge, CO. Rates of chlorothalonil loss from the verdure at Vail from 5 Nov 2006 2 to 26 Mar 2007 were not different (P >0.10) between the high (-0.3 µg/g tissue/day, R = 2 0.62, P =0.02) and low (-0.24 µg/g tissue/day, R =0.86, P =0.002) rates. Similarly chlorothalonil loss rates were not different (P>0.10) between high (-0.2 µg/g tissue/day, 2 2 R = 0.90, P = 0.001) and low rates (-0.09 µg/g tissue/day, R =0.76, P =.0 01) at Breckenridge. Shaded area represents period of snow cover during the winter.

Figure 1.3. Relationship between chlorothalonil and fludioxonil residues in the verdure and sampling dates during the winter of 2006-2007 following application of the commercial fungicide mixture Instrata containing chlorothalonil (3.7 kg/ha), fludioxonil (0.006 kg/ha) and propiconazole (0.093 kg/ha), to an annual bluegrass fairway at Vail and a Kentucky bluegrass fairway at Breckenridge, Colorado. The rate of fludioxonil loss from the verdure 5 Nov 2006 to 26 Mar 2007 were not different (P >0.10) between 2 Vail (-0.004 µg/g tissue/day, R = 0.44, P = 0.09) and Breckenridge (-0.006 µg/g 2 tissue/day, R = 0.65, P = 0.03). Shaded area represents period of snow cover.

Figure 1.4. Relationship between chlorothalonil and fludioxonil residues in the verdure and sampling dates during the winter 2007-08 following application of two rates of the commercial fungicide mixture Instrata containing chlorothalonil (3.0 and 3.7 kg/ha), fludioxonil (0.005 and 0.006 kg/ha) and propiconazole (0.076 and 0.093 kg/ha), and fludioxonil (0.008 kg/ha) applied alone (as the commercial fungicide Medallion 50WP) to an annual bluegrass fairway at Vail, Colorado. Shaded area represents period of permanent snow cover.

Figure 1.5. Inhibition in diameter growth of Typhula ishikariensis isolates on potato dextrose agar amended with 1,10, 100, and 500 µg/ml chlorothalonil relative to growth in non-amended agar. A subset of isolates was screened at the higher concentrations.

Figure 1.6. Inhibition in diameter growth of Typhula incarnata isolates on potato dextrose agar amended with 1, 10, 100, and 500 µg/ml chlorothalonil relative to growth in non-amended agar.

Figure 1.7. Growth inhibition of Typhula ishikariensis on half-strength potato dextrose agar amended with 0.001, 0.01, 0.1 and 1 µg/ml fludioxonil relative to growth of isolates in non-fungicide amended agar.

Figure 1.9. Growth inhibition of Typhula ishikariensis isolates on half strength potato dextrose agar amended with 1 and 10 µg/ml propiconazole relative to growth in non- amended agar. A subset (57) of all isolates (179) was screened at the higher concentration.

Figure 1.10. Growth inhibition of Typhula incarnata isolates on half strength potato dextrose agar amended with 1 and 10 µg/ml propiconazole relative to growth in non- amended agar.

Figure 1.11. Concentration study (2008-09) using chlorothalonil applied at 10X the recommended rate showing that development of snow mold was inhibited; however, phytotoxicity damage to turfgrass was exhibited.

Figure 1.12. Typhula blight damage to A) chlorothalonil treated (8.1 kg/ha), B) chlorothalonil treated (13.9 kg/ha), and C) an untreated plot on a Kentucky bluegrass fairway at Breckenridge, CO in May 2006. Area between the lines is an individual treatment plot.

Figure 1.13. Typhula blight damage to A) chlorothalonil-treated plot (8.1 kg/ha), B) chlorothalonil treated plot (13.9 kg/ha), and C) an untreated plot on an annual bluegrass fairway at Vail, CO in March 2007. Area between the black lines is an individual treatment plot.

Figure 1.14 Representative isolates of Typhula spp. Plate labeled 581 is Typhula incarnata; Plate labeled 411 is Typhula ishikariensis var. canadensis/BSII; Plate labeled 195 is Typhula idahoensis/BSI; and Plate labeled SM2 is Typhula ishikariensis var. ishikariensis/BSI. Note differences in color and size of the sclerotia.

Table 1.1. Typhula blight damage to a Kentucky Bluegrass fairway at Breckenridge, and an annual bluegrass fairway at Vail, CO, following fungicide applications.

Treatment and Rate % of Plot Damagedx

Breckenridge Breckenridge Vail Vail 2005-06 2006-07 2006-07 2007-08 Quintozene 15.12 kg/ha (Turfcide 400) 17.5b 24.0bc 50.0c Chlorothalonil 8.06 kg/ha (Daconil Ultrex 82.5WDG) 58.8a 20.0bc 73.75b Chlorothalonil 13.86 kg/ha 50.0a 40.0b 82.0ab Chlorothalonil 3.7 kg/ha, Propiconazole 0.093 kg/ha, Fludioxonil 0.006 kg/ha (Instrata 3.61SE) 7.5c 48.75c 22.5d Chlorothalonil 2.98 kg/ha, Propiconazole 0.076 kg/ha, Fludioxonil 0.005 kg/ha 36.3cd Fludioxonil 0.765 kg/ha (Medallion 50WP) 75.8b Propiconazole 1.075 kg/ha (BannerMAXX 1.3ME) 52.5c Untreated Control 42.5a 71.25a 87.0a 98.8a xPercent damage is the mean of 4 replications. Means not followed by the same letter in the same column are different (P<0.05) according to a F-least significant difference test.

Table 1.2. Typhula blight damage to an annual bluegrass fairway at Vail, CO, in 2009 following applications of chlorothalonil and fludioxonil at labeled and 2X, 5X and 10X labeled rates.

% of Plot Damaged (means of 4 Treatment and Rate treatments) Labeled Rate Daconil Ultrex Chlorothalonil 82.5WDG 12.51 kg/ha 83.0ab Chlorothalonil 2X 25.22 kg/ha 52.5c Chlorothalonil 5X 63.08 kg/ha 58.8bc Chlorothalonil 10X 126.04 kg/ha 14.3d Untreated Control 94.0a Labeled Rate Medallion 50WP Fludioxonil 0.38 kg/ha 74.3a-c 2X Fludioxonil 0.77 kg/ha 78.0a-c 5X Fludioxonil 1.91 kg/ha 82.5ab 10X Fludioxonil 3.86 kg/ha 62.5bc xValues are means of four replications. Means not followed by the same letter are different (P<0.05) according to an F-least significant difference test. yTurfgrass in plots was extensively discolored as a result of phytotoxicity from the high rate of chlorothalonil.

Table 1.3. Snow mold isolates identified by ITS sequencing. Biological species I is Typhula ishikariensis var. ishikariensis or T. ishikariensis var. idahoensis; Biological species II is T. ishikariensis var. canadensis.

Snow Mold Isolates Sequenced Vail Breckenridge Other Locations Typhula ishikariensis- Biological species I 40 17 16 Typhula ishikariensis var. canadensis- Biological species II 14 7 9 Typhula incarnata 8 0 13 Typhula phacorrhiza 0 0 4 Myriosclerotinia borealis 0 0 1 Microdochium nivale 1 0 0 Waitea circinata 0 0 1

LITERATURE CITED:

Arsvoll, K. and J. D. Smith 1978. Typhula-Ishikariensis And Its Varieties, Var-idahoensis Comb-Nov And Var-canadensis Var-Nov. Canadian Journal of Botany-Revue 56(3): 348-364.

Beard, J. B. 1973. Turfgrass: Science and Culture. Prentice-Hall Publishing, NJ

Bruehl, G. W. and Cunfer, B.M. 1975. Typhula Species Pathogenic To Wheat In Pacific Northwest. Phytopathology 65(7): 755-760.

Bruehl, G. W. and Machtmes, R. 1980. Cultural Variation Within Typhula-idahoensis And Typhula-ishikariensis And The Species Concept. Phytopathology 70(9): 867-871.

Bruehl, G. W., Machtmes,R., and Kiyomoto, R. 1975. Taxonomic Relationships Among Typhula Species As Revealed By Mating Experiments. Phytopathology 65(10): 1108- 1114.

Carroll, M. J., Hill, R.L., Pfeil, E., and Herner, A.E. 1993. Washoff Of Chlorothalonil From Turfgrass Foliage. International Turfgrass Society Research Journal. 7: 964-970.

Carroll, M. J., Hill, R. L., Pfeil, E., and Krouse, J. M. 2001. Effect Of Residence Time On Washoff Of Chlorothalonil From Turf Foliage. International Turfgrass Society Research Journal. 9(Part 1): 1-5.

Chang, S. W., Scheef, E., Abler, R.A.B., Thomson, S., Johnson, P., and Jung, G. 2006. Distribution Of Typhula Spp. And Typhula ishikariensis Varieties In Wisconsin, Utah, Michigan, And Minnesota. Phytopathology 96(9): 926-933.

Couch, H. B. 1995. Diseases of Turfgrass. Krieger Publishing Co., Malabar, FL

Durborow, T. E., Barnes, N.L., Cohen, S.Z., Horst, G.L., and Smith, A.E. 2000. Calibration And Validation Of Runoff And Leaching Models For Turf Pesticides, And Comparison With Monitoring Results. Fate and Management of Turfgrass Chemicals. Amer Chemical Soc. 743: 195-227.

Fletcher, J. S., Nellessen, J.E., and Pfleeger, T.G. 1994. Literature-Review And Evaluation Of The Epa Food-Chain (Kenaga) Nomogram, An Instrument For Estimating Pesticide-Residues On Plants. Environmental Toxicology and Chemistry 13(9): 1383- 1391.

Hsiang, T., Matsumoto, N., and Millet, S.M. 1999. Biology And Management Of Typhula Snow Molds Of Turfgrass. Plant Disease 83(9): 788-798.

Hsiang, T. and Wu, C.R. 2000. Genetic Relationships Of Pathogenic Typhula Species Assessed By RAPD, ITS-RFLP And ITS Sequencing. Mycological Research 104: 16-22.

Jo, Y. K., Niver, A.L., Rimelspach, J.W., and Boehm, M.J. 2006. Fungicide Sensitivity Of Sclerotinia homoeocarpa From Golf Courses In Ohio. Plant Disease 90(6): 807-813.

Mastovska, K. and Lehotay, S.J. 2004. Evaluation Of Common Organic Solvents For Gas Chromatographic Analysis And Stability Of Multiclass Pesticide Residues. Journal of Chromatography A 1040(2): 259-272.

Matsumoto, N., Sato, T. and Araki, T. 1982. Biotype Differentiation In The Typhula ishikariensis Complex And Their Allopatry In Hokkaido. Ann. Phytopath. Soc. Jpn. 48: 275-280.

McBeath, J. H. 2002. Snow Mold-Plant-Antagonist Interactions: Survival Of The Fittest Under The Snow. The Plant Health Instructor.

Monadjemi, S., El Roz, M., Richard, C., and Ter Halle, A. 2011. Photoreduction Of Chlorothalonil Fungicide On Plant Leaf Models. Environmental Science & Technology 45(22): 9582-9589.

Potter, T. L., Wauchope, R.D., and Culbreath, A.K. 2001. Accumulation And Decay Of Chlorothalonil And Selected Metabolites In Surface Soil Following Foliar Application To Peanuts. Environmental Science & Technology 35(13): 2634-2639.

Sigler, W. V., Reicher, Z., Throssel, C., Bishoff, M., and Turco, R.F. 2003. Sorption And Degradation Of Selected Fungicides In The Turfgrass Canopy. Water Air and Soil Pollution 142(1-4): 311-326.

Skoglund, L. 1998. Snow Molds In Colorado; A Review. Unpublished.

Smiley, R. W., Dernoeden, P.H., and Clarke, B.B. 2005. Compendium of Turfgrass Diseases, Third Edition. American Phytopathological Society, St. Paul, MN

Smith, J. D., Jackson, N., and Woolhouse, A.R. 1989. Fungal Diseases of Amenity Turf Grasses., E. & F.N. Spon, New York, NY.

Stromqvist, J. and Jarvis, N. 2005. Sorption, Degradation And Leaching Of The Fungicide Iprodione In A Golf Green Under Scandinavian Conditions: Measurements, Modelling And Risk Assessment. Pest Management Science 61(12): 1168-1178.

Vargas, J. M., Beard, J.B., and Payne, K.T. 1972. Comparative Incidence Of Typhula Blight And Fusarium Patch On 56 Kentucky Bluegrass Cultivars. Plant Disease Reporter 56(1): 32-&.

Vargas, J. M. J. 1994. Management of Turfgrass Diseases. CRC Press, Boca Raton, FL

Wang, G. L., Liang, B., Li, F., and Li, S.P. 2011. Recent Advances In The Biodegradation Of Chlorothalonil. Current Microbiology 63(5): 450-457.

Wang, Z., Casler, M.D., Stier, J.C., Gregos, J.S. and Millett, S.M. 2005. Genotypic Variation For Snow Mold Reaction Among Creeping Bentgrass Clones. Crop Science 45(1): 399-406.

Willis, G. H. and McDowell, L.L. 1987. Pesticide Persistence On Foliage. Reviews of Environmental Contamination and Toxicology 100: 23-73.

Wu, L., Liu, G., Yates, M.V., Green, R.L., Pacheco, P., Gan, J., and Yates, S.R. 2002. Environmental Fate Of Metalaxyl And Chlorothalonil Applied To A Bentgrass Putting Green Under Southern California Climatic Conditions. Pest Management Science 58(4): 335-342.

CHAPTER 2

EFFECT OF SNOW REMOVAL ON GRAY SNOW MOLD DEVELOPMENT AT HIGH

ALTITUDE GOLF COURSES.

SUMMARY

Golf course superintendents at high altitudes in Colorado apply fungicides in late

October before permanent snow cover to prevent gray (Typhula incarnata) or speckled snow mold (Typhula ishikariensis) development. Snow is also removed from putting greens in March and kept snow-free through spring to help suppress snow mold.

However, the benefits of spring snow removal in snow mold suppression compared to potential turfgrass damage caused by exposure to low temperatures following removal have not been documented. We compared snow mold severity and turfgrass health on a Kentucky bluegrass fairway at Breckenridge and annual bluegrass plots at Vail, CO with permanent snow cover through winter to plots where snow was removed from late

October through mid-November and to plots where snow was removed in mid-March and maintained snow-free through the spring. Both gray and speckled snow molds were observed at both locations and in each year, although the majority of the damage was caused by speckled snow mold. Snow removal in the fall resulted in high fluctuation of temperatures at the turfgrass surface until permanent snow cover stabilized them for the remainder of the winter. Snow removal in the spring also resulted in extreme temperature fluctuations, but damage to the turfgrass by freezing

50 temperatures was not apparent. A similar trend in temperature fluctuations was observed for each treatment in each year and at both locations.

Introduction:

Gray snow mold caused by Typhula incarnata and speckled snow mold caused by T. ishikariensis, collectively called Typhula blight, are major diseases of turfgrasses at high elevations in Colorado. These fungi oversummer in the thatch as hardened sclerotia. Scleortia germinate in fall and the mycelium colonizes the above-ground portions of plants. Sclerotia of T. incarnata may also germinate to form sporocarps that produce basidiospores. However, sporocarp formation is apparently restricted to regions with rainy or foggy fall weather (Smith, 1989), and has not been observed in

Colorado. Although turf can be damaged by T. incarnata from late autumn to early spring in wet, cold climates with little to no snow, both T. incarnata and T. ishikariensis are more often active under deep, prolonged snow cover (Smith, 1989). In spring after snowmelt, patches of matted turfgrass with gray mycelium are apparent. In severe cases, coalescing patches may cover the entire sward. Numerous sclerotia are formed on diseased leaf surfaces or are embedded in leaf tissue. Damage from Typhula blight is variable; in some years only the leaves are killed, whereas in others large areas of the turfgrass may be killed.

Typhula blight is primarily controlled with preventive fungicide applications in fall just prior to snow cover. Efficacy can be influenced by the active ingredient used, application timing, and disease severity. Numerous fungicides and fungicide

51 combinations have been tested for Typhula blight control (Koch, 2010a, b, and c, Koch,

2011, Popko, 2011a, b, and c) ( Appendix 2 ). In general, fungicide combinations having different modes of action are recommended with the assumption that these mixtures provide a broader spectrum of control against a variety of snow mold pathogens (Chang et al., 2006, Jo et al., 2006, Jung, 2007, Vargas, 1994). Fungicides are often applied as a single application as close to snow cover as possible to prevent chemical dissipation or photodegradation. For example, the dissipation half-life of chlorothalonil on a bentgrass putting green in the summer was 5 days (Wu et al., 2002).

In another study, the field-extrapolated half-life of chlorothalonil photolysis on vegetation was also estimated to be 5 days (Monadjemi et al., 2011). In contrast, chlorothalonil persisted for up to five months under snow without appreciable dissipation (Chapter 1).

It has been recommended that systemic fungicides should be applied before leaf growth completely stops in the fall to allow for uptake and movement, although details on timing weren’t provided (Smiley, 2005). This premise has been adopted and modified by some

Colorado superintendents into the practice of making two fall applications for Typhula blight control; one fungicide application containing a systemic triazole or strobiluron fungicide in late September or early October, followed by a second application in late

October or early November just prior to snow cover.

Late winter or early spring snow removal is often cited as a control strategy for

Typhula blight (Hsiang et al., 1999, Smiley, 2005). This is based on observations that

Typhula blight is more severe in areas with a longer duration of snow cover (Chang et al., 2006, Hsiang et al., 1999, Smith, 1989). Late winter snow removal from putting

52 greens has been widely implemented by Colorado superintendents as a way to reduce

Typhula blight, yet there is no experimental data to support this practice.

Superintendents also remove snow in late winter to prevent ice or freeze damage to the turfgrass. Winter injury, resulting from low temperatures or ice cover is important in cold climate areas (Beard and Olien, 1963), including Colorado. The injury is often exacerbated if the predominant grass species is annual bluegrass (Poa annua)

(Tompkins et al., 2004), which is the most common species of turfgrass on high elevation golf courses in Colorado. Contrary to the practice of early snow removal,

Tompkins et al. (Tompkins et al., 2000) concluded that it was important to maintain a snow cover on putting greens in order to maintain turfgrass dormancy for as long as possible.

The objectives of my study were to examine the efficacy of various fungicides against Typhula blight, and to determine whether two fall fungicide applications enhanced control relative to a single application just before snow cover. I also determined whether certain cultural practices, including fall and spring snow removal, and snow compaction influenced Typhula blight development.

Methods and Materials:

Effect of fungicide and snow removal on snow mold development 2005-06.

Plots were established on an annual bluegrass golf course fairway at Vail and a

Kentucky bluegrass fairway at Breckenridge, CO on 24 Oct 2005 to determine the

53 effects of fungicide applications and snow removal timing on Typhula blight damage.

Plots were arranged in a split-plot randomized complete-block design with 4 replications. Whole plots measured 1.5 X 4.3 m and were left untreated or treated with quintozene (Turfcide 400F, American Vanguard Corporation, Newport Beach, CA) at

15.12 kg a.i./ha. Applications were made using a CO2 sprayer pressurized to 138 kPa and equipped with a spray boom with four 8004 flat fan nozzles to deliver water at 815 L per ha. Subplots measured 1.4 X 2.1 m and consisted of; no snow removal during the experiment; snow removal at approximately weekly intervals from the application date until the end of November (Fall) and thereafter allowing snow to accumulate on plots until spring melt, or; snow removal at approximately weekly intervals from 1 Mar 2006 until the final rating date on 27 Apr 2006 (Spring). Snow was removed with shovels

(Figure 2.3). Snow covered the plots on 21 Nov 2005 and remained until 20 Apr 2006.

Temperatures at the turfgrass surface in each snow-removal treatment were recorded at two hour intervals for the duration of the experiment using thermistors (Spectrum

Technologies, Plainfield, IL).

Typhula blight severity was determined at the last sampling date on 7 May 2006 by visually estimating the percent damage in each plot. Data were subjected to analysis of variance and means separated using Fisher’s protected least significant difference

(LSD) (P=0.05). Typhula blight severity in the non-fungicide, non-snow removal subplots was also collected at approximately monthly intervals through the winter. On the first and last sampling dates with no snow cover, sub-plots were rated visually.

However, Typhula blight severity in plots on other dates with snow cover were estimated by arbitrarily removing two, 6.35-cm-diameter X 5-cm-deep turf cores from

54 each plot using a 1.2-meter-long soil core remover (Figure 2.4) and determining percentage of dead grass in each core. The percent damage in the two cores from each subplot were averaged; those percentages were then used to determine the mean snow mold damage among the four replicates in each treatment.

The experiment was repeated at Vail and Breckenridge in 2006-2007 using the same plot design as previously described. The commercial fungicide mixture Instrata

3.6SE (Syngenta Professional Products, Greensboro, NC) containing 3.7 kg chlorothalonil, 0.006 kg fludioxonil and 0.093 kg propiconazole a.i. per ha was applied at

Vail, while quintozene (Turfcide 400F, American Vanguard Corporation, Newport

Beach, CA) at 15.12 kg a.i./ha was applied at Breckenridge. Fungicide applications were made on 24 Oct 2006 at both locations and final snow mold damage ratings were made on 1 May 2007. Temperature measurements and turfgrass core collections also were the same as in 2005-06, except that cores were placed in a layer of moistened sand in 30-cm-diameter plastic pots and incubated in the greenhouse for two weeks after which they were rated for percentage turf damage (Figure 2.5).

A third experiment was conducted at Vail in 2007-08. Plot sizes were the same as in 2005-06. On 24 Oct 2007 the commercial fungicide mixture Instrata 3.6SE

(Syngenta Professional Products, Greensboro, NC) at the same application rates of

2006-2007 was applied to whole plots. Subplots consisted of no snow removal or a spring removal as previously described. A fall snow removal treatment was not included and turf cores were not collected during the winter. The final plot rating was on

12 May 2008.

Effect of snow compaction on temperatures at the turfgrass surface

Nordic track ski trails were created for cross-country skiing and snowshoeing at the Breckenridge and Vail golf courses by compacting snow covering the fairways and tee boxes using a track-driven vehicle (snow cat). Trails were groomed and compacted following each snow event during the winter. The level of snow compaction was not measured. Temperatures at the snow/turfgrass interface beneath the Nordic track and an adjacent non-compacted area on a golf course fairway at each course were measured at 2-hour intervals with thermistors (Spectrum Technologies, Plainfield, IL) throughout the winter in 2006-07 and again in 2007-08.

Fungicide efficacy trials

Fungicide efficacy trials have been conducted each year since 1984 to determine which chemical/biological treatments provide the best control of Typhula blight.

Fungicides were applied to a Kentucky bluegrass fairway at Breckenridge or to an annual bluegrass fairway or putting green at Vail, CO. Treatments were arranged in a randomized complete block with three to four replications. Individual plots measured

1.5 x 1.5 m on the fairways and 0.9 x 0.9 m on the putting green. Fungicides were applied in late October to early November with a CO2 powered sprayer as previously described: granular formulations were spread using a hand-held shaker. In some cases sand was added to the granular formulations to increase volume and allow for more uniform coverage of the plot during application. Plots were rated in April or May

56 after snow melt for percentage turfgrass damage by Typhula blight. Methods and results of these studies from 2005-2011 are presented in Appendix 2.

In addition to comparing the efficacy of different fungicide products, studies were conducted to determine whether two fall fungicide applications were superior to a single application just before snow cover in controlling Typhula blight. In these studies, a combination of fungicides were applied in late September or early October approximately one month before snow cover. A second application of these same fungicide combinations, or in some cases a slightly different fungicide combination, was applied in late October (Table 2.1). These treatments were compared by paired T-tests to plots treated with the same fungicide combinations applied in late October only

(Table 2.1).

Results

Effect of fungicide and snow removal on snow mold development

Both gray and speckled snow molds were observed at both locations and in each year, although the majority of the damage was caused by speckled snow mold. Pink snow mold was not common (<1% of the patches) in any year.

Typhula blight severity was consistently higher on an annual bluegrass fairway at

Vail compared to a Kentucky bluegrass fairway at Breckenridge (Figure 2.6).

Applications of quintozene at Vail and Breckenridge in 2005-2006 and 2006-2007, and the formulated product Instrata at Vail in 2007-08 (Figure 2.10) reduced (P<0.05), but

57 did not completely suppress Typhula blight (Figure 2.6 and 2.7). There were no interactions in Typhula blight damage between fungicide and snow removal treatments, except at Vail (P<0.5) in 2005-06 where severe damage (86%) was observed in the fungicide-treated plots that had snow removed in the fall. Most of the damage was a result of freeze injury and not Typhula blight as determined by the lack of mycelium and sclerotia in the dead grass. Fall snow removal did not increase winter damage or

Typhula blight in Kentucky bluegrass at Breckenridge in either year (Figure 2.6 and 2.7).

Spring snow removal also did not reduce (P>0.10) snow mold severity in fungicide- treated or non-treated subplots compared to subplots with no snow removal.

Snow removal treatments influenced temperatures at the turfgrass surface during the winter; a similar trend was observed for each treatment in each year and at both locations (Figure 2.11, 2.12 and 2.13). In the no-snow removal plots, surface temperatures fluctuated prior to snow cover, and then stabilized between -1°C and 0°C for the duration of winter until snow melted in spring. In the fall snow removal plots, temperatures fluctuated widely, with temperatures reaching lows of -18°C on some days in late November 2005 at Vail (Figure 2.11). Similar trends were seen in other years

(Figure 2.12 and 2.13). After snow cover, temperatures slowly increased and then stabilized by mid-December to near freezing for the remainder of the winter. The freezing and thawing cycles in late November resulted in increased damage to the annual bluegrass at Vail, but not Kentucky bluegrass at Breckenridge. Temperatures in the spring-snow removal plots mimicked those in the no-snow removal plots from

November until March. Then temperatures at the turfgrass surface fluctuated widely, and multiple freezing and thawing cycles occurred after snow removal. Yet no

58 increased turf damage was observed as a result of fluctuating temperatures in the spring-snow removal plots (Figure 2.6, 2.7 and 2.10). Maximum temperatures recorded at the surface during periods of no snow cover were likely higher than what actually occurred because the thermistors may have been heated by direct exposure to sunlight.

Temporal development of Typhula blight was monitored in the no-snow removal plots in 2006-07 and 2007-08 at Vail and Breckenridge (Figure 2.8 and 2.9). No mycelium or sclerotia of Typhula spp. were observed on cores removed during

November in either year. On 23 Dec 2006, a very sparse mycelium of Typhula

(determined by microscopic confirmation of clamp connections) was observed on cores collected at Vail and Breckenridge, however, turf necrosis was not observed. Similarly, no turf damage was observed on 6 Dec 2007 at either location. However, Typhula blight severity had increased dramatically at both locations and in both years by the

January sampling period (Figure 2.8 and 2.9). In fact, the disease severity recorded in

January, with the exception of Vail in 2007-08, was similar to the final severity ratings taken in April or May.

Effect of snow compaction on temperatures at the turfgrass surface

In 2005-06, Typhula blight was observed in a non-snow compacted area adjacent to, but not under, a snow-compacted Nordic ski track path that had been established on the Kentucky bluegrass fairways and tees at Breckenridge. This phenomenon was observed again in 2006-07 (Figure 2.14 and 2.15). Temperatures at the turfgrass surface in the non-compacted areas remained near 0°C, whereas

59 temperatures under the compacted areas were continuously below freezing and consistently 3-7°C colder during the majority of the winter (Figure 2.16 and 2.17).

Although the turfgrass under the Nordic track exhibited few Typhula blight patches following snow melt, it was discolored and took several weeks longer than the non- compacted areas to resume growth in the spring (Figure 2.14 and 2.15).

Fungicide Efficacy Trials

Results of the fungicide efficacy trials from 2005-06 through 2010-11 are included in Appendix 2. Although a detailed analysis of each experiment is not provided here, some general observations can be made. Typhula blight damage (>90% each year) on an untreated annual bluegrass fairway at Vail was always more severe than a

Kentucky bluegrass fairway at Breckenridge. Many of the treatments provided only fair to poor control at Vail, although some resulted in adequate (<10% damage) suppression of Typhula blight (Appendix 2). Several biological control products were tested, but none provided satisfactory control of Typhula blight.

One of the key comparisons in each year of the study was the efficacy of a late-

September application of a fungicide combination followed by a second application in late October, compared to a single fungicide application prior to snow cover (Table 2.1).

Twenty paired comparisons from two golf courses over a 7-year period were analyzed and in only one case did the two application regime provide better (P<0.05) control of

Typhula blight than the single fungicide application.

Discussion

Kentucky bluegrass and annual bluegrass have been reported as intermediate in susceptibility to T. incarnata and T. ishikariensis (Wu and Hsiang, 1998). In my studies, annual bluegrass was consistently more severely damaged by Typhula blight than

Kentucky bluegrass. Not only were the patches less numerous in Kentucky bluegrass, they were more superficial and the turfgrass appeared to recover quickly once the turfgrass resumed growth. Golf course superintendents in Colorado that manage

Kentucky bluegrass fairways and that have limited fungicide budgets may be able to forego fall fungicide applications without suffering significant turfgrass damage from

Typhula blight.

Golf course superintendents in the Colorado Mountains routinely remove snow from putting greens beginning in late February and keep the greens snow-free for the duration of the winter. Many do this in the belief that it helps suppress snow mold development. Snow molds are reported to be more severe during winters with extended periods of snow cover (Hsiang et al., 1999; McBeath, 2002; Nissinen, 1996) and snow removal is often recommended as a control (Hsiang et al., 1999; Smiley,

2005; Smith, 1989). Repeat fungicide applications after a mid-winter thaw or snow removal also have been suggested (Burpee et al., 1990; Fushtey, 1980; Smiley, 2005).

However, in these experiments, Typhula blight severity at the time of snow removal in early March in the non-fungicide treated subplots was very similar to that observed in the no-snow removal plots rated in late April or May. For example, the non-fungicide treated annual bluegrass subplots had already been damaged >90% by Typhula blight

61 at the time of snow removal in March. Residue studies (Chapter 1) also indicated that fungicide concentrations in the verdure did not appreciably decline from November through March, thus obviating the need to re-treat the turfgrass after snow removal.

Therefore, this research did not support the practice of late winter snow removal in the early spring to help suppress Typhula blight development on fungicide-treated turfgrass.

Another reason superintendent’s remove snow from putting greens in late winter is to prevent ice formation resulting from melting snow under snowpack. The combination of crown hydration followed by ice formation is a major cause of winter damage to annual bluegrass in some regions (Dionne et al., 2001; Fry, 2004; Tompkins et al., 2000, 2004). There was no ice formation or freeze injury to annual bluegrass or

Kentucky bluegrass in the no-snow removal subplots in any year of these studies. Turf quality was similar to that in the spring snow removal plots. Thus, my results did not support the need for early snow removal to prevent freeze injury. However, plots were established on a level fairway rather than a putting green and it is possible that the higher mowing height may have protected crowns from freeze injury. Furthermore, ice damage may be an issue in low areas of fairways or putting greens where water is more likely to puddle and freeze.

Late winter snow removal can expose turfgrass to extreme temperature fluctuations and multiple subfreezing events as evidenced by temperature data in the snow removal subplots. Annual bluegrass quality immediately following snow removal in early March was excellent in all subplots; surprisingly there was no evidence of ice formation or freeze injury to the turfgrass in any year. The turfgrass in the spring snow removal subplots continued to remain healthy throughout the spring even though it was

62 exposed to multiple freezing/thawing events. Cold-acclimated plants are more tolerant of freezing temperatures (Fry, 2004), and it is possible that the extended exposure of turf under snow cover led to a high level of cold-acclimation.

The hypothesis was that snow removal in late fall would freeze the upper soil profile and result in surface temperatures during the winter too cold for snow mold development. Although Typhula spp. are psychrophilic, their growth is inhibited when soil temperatures drop below 0°C (Cunfer and Bruehl, 1973; Dejardin and Ward, 1971;

Hoshino et al., 1997; McBeath, 2002.). In these plots, snow removal in November resulted in temperatures fluctuating between a low of 15-18°C and a high of 25°C at the turfgrass surface until permanent snow cover provided temperature stabilization.

However, temperatures eventually increased and remained near 0°C, and similar to the no-snow removal plots, throughout the winter. Therefore there was no advantage, in terms of reducing surface temperatures below freezing for long periods, with this treatment. However, fall snow removal did result in significant freeze damage to the annual but not Kentucky bluegrass in 2005-06. Golf course superintendents in

Colorado often apply nitrogen fertilizers and continue to irrigate late into fall to encourage turfgrass recovery and to promote golf play for as long as possible. These practices may inhibit cold-acclimation and limit winter hardiness (Fry, 2004; Smith,

1989).

Typhula sclerotia germinate in the soil or thatch during cool, wet weather just prior to or immediately following snow cover to produce mycelium (Hsiang et al., 1999;

Vargas, 1994). The mycelium then colonizes plant tissue directly or through stomata

(Oshiman et al., 1995), although the rate at which this occurs is variable depending on

63 environmental conditions. Although mycelial growth apparently begins soon after snow cover in late fall, long snow durations are required for plant colonization and significant plant damage (Hsiang et al., 1999; Smith, 1989; Vargas, 1994). The rate at which these fungi colonize tissue has not been thoroughly studied. McBeath (2002.) reported that maximum snow mold activity in Alaska occurred in late spring as the snow began to melt and when soil temperatures were near 0°C and the soil/snow interface was saturated. These same conditions were observed soon after snow cover in late fall and in the absence of snow melt. Yet there was no evidence of mycelial growth or plant colonization in November in cores removed under snow cover, and only sparse fungal growth and no plant damage in December. By early to mid-January, extensive mycelial growth on plants was observed, suggesting that plant colonization occurred over a relatively short time period. Whether the turfgrass had been colonized by Typhula at a level not visually observable is not known, but these results suggest plant colonization is not necessarily a gradual process.

Preventing snow compaction has been offered as a management tool for suppressing Typhula blight (Smiley, 2005). Snow compaction may prevent rapid snow melting in spring and could result in longer periods of snow cover. However, it was noted that snow compaction lowered the surface temperature to a level that is inhibitory to Typhula growth (Hsiang et al., 1999; McBeath, 2002;, Matsumoto, 2009; Smith,

1989,). These data were corroborated by the lack of Typhula blight development under the Nordic track. Although snow compaction could be an effective strategy for reducing

Typhula blight, its application may have other detrimental effects. For example, annual and Kentucky bluegrass in compacted areas exhibited more discoloration and a slower

64 spring greenup relative to non-compacted areas. Compaction also resulted in irrigation line damage at Breckenridge in 2006 as a result of freezing pipes. Thus compaction may be of limited use on golf courses for control of snow molds caused by Typhula.

Another control strategy for Typhula blight that is of questionable value in

Colorado, but isstill used, is the practice of making a fungicide application in late

September or early October followed by a second application just before snow cover.

The origin of this concept is unknown, although it may be based on the belief that sclerotia of Typhula germinate during wet, cool periods in the fall to form basidiospores that are subsequently dispersed by air currents. Fungicide applications presumably prevent this early inoculum buildup prior to snow cover. However, sporocarps of T. ishikariensis are rare in nature (Cunfer and Bruehl, 1973) and have never been observed in Colorado. In addition, it is assumed that uptake and movement of systemic fungicides is better when the turfgrass is actively growing. In my studies, early fall fungicide applications, with one exception, did not enhance Typhula blight control. The significant expense of this early fungicide application cannot be justified based on these results.

Figure 2.1. Golf Course superintendents at high elevation courses in Colorado routinely remove snow from putting greens in late February or early March.

Figure 2.2. Annual bluegrass putting green after snow removal in late February 2006. Note that there is no discoloration or evidence of freeze injury to the turfgrass.

Figure 2.3. Snow removal from plot in March 2008.

Figure 2.4. Removal of soil cores with a 1.2-meter-long soil core remover during 2005- 06.

Figure 2.5. Example of gray and speckled snow mold damage to turfgrass cores removed from snow-covered plots and incubated in greenhouse for 2 weeks (2006-07).

Figure 2.6. Effect of snow removal and fungicide applications on turfgrass damage at Vail and Breckenridge, CO 2005-06. The fungicide quintozene was applied 25 Oct prior to permanent snow cover on 21 Nov 2005. Snow was not removed (none), removed at weekly intervals during November (Fall), or removed at weekly intervals from 5 Mar, 2007 (Spring) until the final rating date on 7 May. Damage resulting from the November snow removal was a combination of winter injury and snow mold development. Means not followed by the same letter are significantly different (P =0.05) by Fisher’s LSD test.

Figure 2.7. Effect of snow removal and fungicide applications on turfgrass damage at Vail and Breckenridge, CO 2006-07. The fungicide quintozene was applied 25 Oct prior to permanent snow cover on 7 Nov 2006. Snow was not removed at weekly intervals from plots (None), removed from 7 Nov to approximately the last week in November (Fall removal), or removed on 5 March, 2007 (Spring removal) until the final rating date on 7 May (presented here). Damage resulting from the November snow removal was a combination of winter injury and snow mold development. Means not followed by the same letter are significantly different (P =0.05).

Figure 2.8. Seasonal development of gray and speckled snow mold in non-fungicide treated plots on an annual bluegrass and Kentucky bluegrass fairway at Vail and Breckenridge, CO respectively in 2005-06. Shaded area represents the period of snow coverage on plots. Snow mold damage on all dates except 27 Apr 2006 were determined by removing turf cores from underneath the snow cover and estimating plot damage based on injury to the cores. Bars for each mean represent standard error.

Figure 2.9. Seasonal development of gray and speckled snow mold in non-fungicide treated plots on an annual bluegrass and Kentucky bluegrass fairway at Vail and Breckenridge, CO, respectively, in 2006-07. Shaded area represents the period of snow coverage on plots. Snow mold damage on all dates except 1 Apr were determined by removing turf cores from underneath the snow cover and estimating plot damage based on injury to the cores. Bars for each mean represent standard error.

Figure 2.10. Effect of snow removal and fungicide applications on turfgrass damage at Vail, CO 2007-08. The commercial fungicide Instrata (containing the active ingredients chlorothalonil, propiconazole and fludioxonil) was applied 25 Oct 2006 prior to permanent snow cover on 7 Nov. Snow was not removed from plots (None), or removed on 26 Mar 2008 (Spring removal) and weekly thereafter until the final rating date 24 Apr 2008. Means not followed by the same letter are significantly different (P =0.05) by Fisher’s LSD test.

Figure 2.11. Temperatures at the turf surface on Kentucky bluegrass fairway at Breckenridge, CO during the winter 2005-06 for plots with no, fall (turquois), spring (purple), and no (brown and green) snow removal. Shaded area represents the period of snow cover.

Figure 2.12. Temperatures at the turf surface on annual bluegrass fairway at Vail, CO during the winter 2005-2006 for plots with no, fall (turquois), and spring (purple) snow removal.

Figure 2.13. Temperatures at the turf surface on annual bluegrass fairway at Breckenridge, CO during the winter 2006-07 for plots with A) fall (brown line) and no snow removal (blue line) and B) spring (blue line) and no snow removal (brown line). The spring and fall snow removal bars indicate the period in which snow was physically removed from the plots at approximately weekly intervals. Shaded area represents the period of snow cover.

Figure 2.14. Snow mold damage was less severe on a Kentucky bluegrass tee at Breckenridge, CO where snow was compacted for Nordic track.

Figure 2.15. Snow mold damage was less severe on a Kentucky bluegrass fairway at Breckenridge, CO where snow was compacted for Nordic track. Note turfgrass discoloration associated with snow compaction.

Figure 2. 16 Temperatures at the surface of Kentucky bluegrass in adjacent areas of a fairway where snow had not been compacted or had been compacted for the construction of a Nordic ski track at Breckenridge CO, 2006-07.

Figure 2.17. Temperatures at the surface of annual bluegrass in adjacent areas of a fairway where snow had not been compacted or had been compacted for the construction of a Nordic ski track at Vail CO, 2006-07.

Table 2. 1 Comparison of one versus two fall fungicide applications for the control of Typhula blight at Vail and Breckenridge, CO 2005-2011.

LocationW, year, treatmentX,Y and rate Percent Damage (± SE)Z Breckenridge 2004-05 BannerMAXX 1.3MEC 4.0 fl oz followed by BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz 11.7 ±9.4 BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 7.5 ±2.5 NS oz

Spectro 90WDG 4.0 oz followed by Spectro 90WDG 4.0 oz + CL-EXP-4 1.0 oz 3.0 ±1.1 Spectro 90WDG 5.76 oz + CL-EXP-4 1.0 oz 2.0 ±1.0 NS

Spectro 90WDG 4.0 oz followed by Spectro 90WDG 4.0 oz + Endorse 2.5WP 4.0 oz 11.8 ±6.2 Spectro 90WDG 5.76 oz + Endorse 2.5WP 4.0 oz 9.0 ±4.1 NS

Untreated control 61.3 ±10.9

Vail 2004-05 BannerMAXX 1.3MEC 4.0 fl oz followed by BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz 25.0 ±2.9 BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 23.7 ±4.2 NS oz

Spectro 90WDG 4.0 oz followed by Spectro 90WDG 4.0 oz + CL-EXP-4 1.0 oz 3.8 ±2.4 Spectro 90WDG 5.76 oz + CL-EXP-4 1.0 oz 4.3 ±2.1 NS

Spectro 90WDG 4.0 oz followed by Spectro 90WDG 4.0 oz + Endorse 2.5WP 4.0 oz 6.3 ±1.3 Spectro 90WDG 5.76 oz + Endorse 2.5WP 4.0 oz 10.0 ±2.0 NS

Untreated control 93.8 ±4.7

Breckenridge 2005-06 BannerMAXX 1.3MEC 4.0 fl oz followed by BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz 11.3 ±5.5 BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 27.8 ±17.7 NS oz

Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6SE 5.5 fl oz 1.0 ±0.7 Instrata 3.6SE 11.0 fl oz 0.0 NS

Untreated control 38.8 ±12.1

Vail 2005-06 BannerMAXX 1.3MEC 4.0 fl oz followed by BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz 25.0 ±6.4 BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 33.7 ±5.9 NS oz

Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6SE 5.5 fl oz 20.5 ±6.1 Instrata 3.6SE 11.0 fl oz 30.0 ±13.3 NS

Untreated control 65.8 ±14.3

Vail 2006-07 Spectro 90WDG 4.0 oz followed by Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz 20.8 ±13.1 Spectro 90WDG 4.0 oz + Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz 33.8 ±16.2 *

Untreated control 88.8 ±3.8

Vail 2007-08 Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6SE 5.5 fl 27.5 ±7.5 oz Instrata 3.6SE 11.0 fl oz 14.3 ±6.6 NS

Untreated control 98.3 ±1.2

Vail 2008-09 Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6SE 5.5 fl oz 23.3 4.8 Fairway 15.0 ±3.5 NS Practice green Instrata 3.6SE 11.0 fl oz Fairway 16.7 ±5.9 NS Practice green 18.3 ±6.6 NS

Untreated control Fairway 92.3 ±5.9 Practice green 98.3

Vail 2009-10 Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6SE 5.5 fl oz

10.0 ±2.9 Instrata 3.6SE 11.0 fl oz 11.7 ±1.7 NS

Curalan 50EG 1.0 oz + Daconil Ultrex 82.5WDG 3.2 oz followed by Insignia 20WG 0.7 oz + Trinity 19.2SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz 23.3 ±1.7 Insignia 20SC 0.54 oz + Trinity 19.2SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz 16.7 ±3.3 NS

Untreated control 81.7 ±1.7

Vail 2010-11 Instrata 3.6SE 5.5 fl oz followed by Instrata 3.6 SE 5.5 fl 0.8 ±0.3 oz Instrata 3.6 SE 11.0 fl oz 2.5 ±1.2 NS

Pegasus HPX 2.75 fl oz + Kestrel MEX 1.3ME 2.0 fl oz + Dovetail 39.3EC 4.0 fl oz + Disarm SC 0.18 fl oz followed by Pegasus HPX 2.75 fl oz + Kestrel MEX 1.3ME 2.0 fl oz + Dovetail 39.3EC 4.0 fl oz + Disarm SC 0.18 fl oz 0.0 Pegasus HPX 5.5 fl oz + Kestrel MEX 1.3ME 4.0 fl oz + Dovetail 39.3EC 8.0 fl oz + Disarm SC 0.36 fl oz 4.0 ±3.7 NS

Interface 24.5SC 3.0 fl oz + Triton Flo 367SC 0.5 fl oz followed by Interface 24.5SC 3.0 fl oz + Triton Flo 367SC 6.3 ±3.8 0.5 fl oz Interface 24.5SC 6.0 fl oz + Triton Flo 367SC 0.85 fl oz 0.0 NS

Untreated Control 90.0 ±0.0 WPlots were established on a Kentucky bluegrass fairway at Breckenridge and at an annual bluegrass fairway at Vail except in 2008-09, when plots were placed on both an annual bluegrass fairway and practice putting green at Vail.

XTreatments with a fungicide application made in late September and followed by a second application in late October or early November just before snow cover were compared to the same or similar fungicide combination applied only at the second date.

YFungicide treatments were propiconazole 14.% a.i. (Banner MAXX and Kestrel MEX), fludioxonil 50.0% a.i (Medallion 50WP), polyoxin D Zinc Salt 2.5% a.i. (Endorse 2.5WP), chlorothalonil 82.5% a.i. (Daconil Ultrex 82.5WDG), chlorothalonil 54.0% a.i. (Pegasus HPX), vinclozolin 50.0% a.i. (Curalan 50EG), pyraclostrobin 20.0% a.i. (Insignia 20WG), triticonazole 19.2% a.i. (Trinity 19.2SC), fluoxastrobin 40.3% a.i. (Disarm 480SC), triticonazole 30.1% a.i. (Triton Flo), chlorothalonil 72% a.i. and thiophanate-methyl 18% a.i. (Spectro 90WDG), iprodione 19.65% a.i. and thiphanate-methyl 19.65% a.i. (Cleary

26/36 and Dovetail EC), iprodione 23.1% a.i. and trifloxystrobin 1.44% a.i. (Interface), chlorothalonil 29.9%, propiconazole 4.7% and fludioxonil 1.2% (Instrata 3.61SE).

Z Damage ratings are the means of 4 replications. A paired T test was used to compare the single fall fungicide application to two fall applications. A * indicates a significant (P<0.05) and NS a non-significant (P>0.10) difference between treatments. Damage from the untreated control plots was included as a reference of overall snow mold severity (pressure) in the experiment.

LITERATURE CITED:

Beard, J.B., and Olien, C.R. 1963. Low Temperature Injury In The Lower Portion Of Poa annua L. Crowns. Crop Sci. 3, 362-3.

Burpee, L.L,, Mueller, A.E., and Hannusch, D.J. 1990. Control Of Typhula Blight And Pink Snow Mold Of Creeping Bentgrass And Residual Suppression Of Dollarspot By Triadimefon And Propiconazole. Plant Disease 74, 687-9.

Cunfer, B.M., and Bruehl, G.W. 1973. Role Of Basidiospores As Propagules And Observations On Sporophores Of Typhula idahoensis. Phytopathology 63, 115-20.

Dejardin, R.A., and Ward, E.W.B. 1971. Growth And Respiration Of Psychrophilic Species Of Genus Typhula. Canadian Journal of Botany 49, 339-&.

Dionne, J., Castonguay, Y., Nadeau, P., and Desjardins, Y. 2001. Freezing Tolerance And Carbohydrate Changes During Cold Acclimation Of Green-Type Annual Bluegrass (Poa annua L.) Ecotypes. Crop Science 41, 443-51.

Fry, J. 2004. Applied turfgrass science and physiology. J. Wiley. Hoboken, NJ

Fushtey, S.G. 1980. Chemical Control Of Snow Mold In Bentgrass Turf In Southern Ontario. Canadian Plant Disease Survey 60, 25-&.

Hoshino, T., Tronsmo, A.M., Matsumoto, N., Ohgiya, S., and Ishizaki, K. 1997. Effects Of Temperature On Growth And Intracellular Proteins Of Norwegian Typhula ishikariensis Isolates. Acta Agriculturae Scandinavica Section B-Soil and Plant Science 47, 185-9.

Hsiang, T., Matsumoto, N., and Millett, S.M. 1999. Biology And Management Of Typhula Snow Molds Of Turfgrass. Plant Disease 83, 788-98.

Koch, P.L. and Kerns, J. P. 2011. Preventative Fungicide Applications For The Control Of Snow Mold On Creeping Bentgrass, 2010-2011. In. Plant Disease Management Reports. (6:T010.)

Koch, P.L. and Kerns, J.P. 2010a. Preventative Fungicide Applications For The Control Of Snow Mold On Creeping Bentgrass, 2009-2010 (I). In. Plant Disease Management Reports. (5:5031.)

Koch, P.L. and Kerns, J.P. 2010b. Preventative Fungicide Applications For The Control Of Snow Mold On Creeping Bentgrass, 2009-2010 (II). In. Plant Disease Management Reports. (5:T032.)

Koch, P.L. and Kerns, J.P. 2010c. Preventative Fungicide Applications For The Control Of Snow Mold On Creeping Bentgrass, 2009-2010 (III). In. Plant Disease Management Reports. (5:T033.)

Matsumoto, N. 2009. Snow Molds: A Group Of Fungi That Prevail Under Snow. Microbes and Environments 24, 14-20.

Mcbeath, J.H. 2002. Snow Mold-Plant-Antagonist Interactions: Survival Of The Fittest Under The Snow. The Plant Health Instructor.

Monadjemi, S., El Roz, M., Richard, C., and Ter Halle, A. 2011. Photoreduction Of Chlorothalonil Fungicide On Plant Leaf Models. Environmental Science & Technology 45, 9582-9

Nissinen, O. 1996. Analyses Of Climatic Factors Affecting Snow Mould Injury In First- Year Timothy (Phleum pratense L.) With Special Reference To Sclerotinia borealis. Acta universitatis Ouluensis. A; Scientiae rerum naturalium 289, 115.

Oshiman, K.I., Kobayashi, I., Shigemitsu, H., and Kunoh, H. 1995. Studies On Turfgrass Snow Mold Caused By Typhula ishikariensis. II. Microscopical Observation Of Infected Bentgrass Leaves. Mycoscience 36, 179-85.

Popko, J., Campbell-Nelson, K., Ok, C. H., and Jung, G. 2011a. Snow Mold Control On A Creeping Bentgrass And Annual Bluegrass Fairway, 2010-2011 (II). In. Plant Disease Management Reports. (6:T028.)

Popko, J., Campbell-Nelson, K., Ok, C. H., and Jung, G. 2011b. Snow Mold Control On A Creeping Bentgrass, Annual Bluegrass And Kentucky Bluegrass Fairway, 2010-2011 (I). In. Plant Disease Management Reports. (6:T029.)

Popko, J., Campbell-Nelson, K., Ok, C. H., and Jung, G. 2011c. Snow Mold Control On A Creeping Bentgrass, Annual Bluegrass And Kentucky Bluegrass Fairway, 2010-2011 (III). In. Plant Disease Management Reports. (6:T027.)

Smiley, R.W., Dernoeden, P.H., and Clarke, B.B. 2005. Compendium of Turfgrass Diseases, Third Edition. St. Paul, MN: American Phytopathological Society.

Smith, J.D., Jackson, N., and Woolhouse, A.R. 1989. Fungal Diseases of Amenity Turf Grasses. New York, NY: E. & F.N. Spon.

Tompkins, D.K., Ross, J.B., and Moroz, D.L. 2000. Dehardening Of Annual Bluegrass And Creeping Bentgrass During Late Winter And Early Spring. Agronomy Journal 92, 5- 9.

Tompkins, D.K., Ross, J.B., and Moroz, D.L. 2004. Effects Of Ice Cover On Annual Bluegrass And Creeping Bentgrass Putting Greens. Crop Science 44, 2175-9.

APPENDIX 1

LIST OF SNOW MOLD ISOLATES

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM101 Vail, CO 5-Mar-07 TISH SM147 Loveland, CO No Date TISH SM102 Vail, CO 5-Mar-07 TISH SM148 Vail, CO 26-Mar-07 TISH SM103 Vail, CO 5-Mar-07 TISH SM149 Vail, CO 26-Mar-07 TISH SM104 Vail, CO 5-Mar-07 TISH SM150 Vail, CO 26-Mar-07 TISH SM105 Vail, CO 5-Mar-07 TISH SM151 Vail, CO 26-Mar-07 TISH SM106 Vail, CO 5-Mar-07 TISH SM152 Vail, CO 26-Mar-07 TISH SM107 Vail, CO 5-Mar-07 TISH SM153 Vail, CO 26-Mar-07 TISH SM108 Vail, CO 5-Mar-07 TISH SM154 Vail, CO 26-Mar-07 TISH SM109 Vail, CO 5-Mar-07 TISH SM155 Vail, CO 26-Mar-07 TISH SM110 Fort Collins, CO No Date TINC SM156 Vail, CO 26-Mar-07 TISH SM111 Fort Collins, CO No Date TINC SM157 Vail, CO 26-Mar-07 TISH SM112 Fort Collins, CO No Date TINC SM158 Vail, CO 26-Mar-07 TISH SM113 Fort Collins, CO No Date TINC SM159 Vail, CO 26-Mar-07 TISH SM114 Fort Collins, CO No Date TINC SM160 Vail, CO 26-Mar-07 TISH SM115 Fort Collins, CO No Date TINC SM161 Vail, CO 26-Mar-07 TISH SM116 Fort Collins, CO No Date TPHA SM162 Vail, CO 26-Mar-07 TISH SM117 Fort Collins, CO No Date TINC SM163 Vail, CO 26-Mar-07 TISH SM118 Fort Collins, CO No Date TINC SM164 Vail, CO 26-Mar-07 TISH SM119 Fort Collins, CO No Date TINC SM165 Vail, CO 26-Mar-07 TISH SM120 Fort Collins, CO No Date TINC SM166 Vail, CO 26-Mar-07 TISH SM121 Fort Collins, CO No Date TINC SM167 Vail, CO 26-Mar-07 TISH SM122 Loveland, CO No Date TISH SM168 Vail, CO 26-Mar-07 TISH SM123 Fort Collins, CO No Date TISH SM169 Vail, CO 26-Mar-07 TISH SM124 Fort Collins, CO No Date TISH SM170 Vail, CO 26-Mar-07 TISH SM125 Loveland, CO No Date TISH SM171 Vail, CO 26-Mar-07 TISH SM126 Fort Collins, CO No Date TISH SM172 Vail, CO 26-Mar-07 TISH SM127 Fort Collins, CO No Date TISH SM173 Vail, CO 26-Mar-07 TISH SM128 Fort Collins, CO No Date TISH SM174 Vail, CO 26-Mar-07 TISH SM129 Loveland, CO No Date TINC SM175 Vail, CO 26-Mar-07 TISH SM130 Loveland, CO No Date TINC SM176 Vail, CO 26-Mar-07 TISH SM131 Loveland, CO No Date TINC SM177 Loveland, CO No Date TISH SM132 Loveland, CO No Date TINC SM178 Loveland, CO No Date TISH SM133 Loveland, CO No Date TINC SM179 Loveland, CO No Date TISH SM134 Loveland, CO No Date TINC SM180 Fort Collins, CO No Date TISH SM135 Loveland, CO No Date TINC SM181 Vail, CO 26-Mar-07 TISH SM136 Fort Collins, CO No Date TINC SM182 Vail, CO 26-Mar-07 TISH SM137 Loveland, CO No Date TINC SM183 Vail, CO 26-Mar-07 TISH SM138 Fort Collins, CO No Date TISH SM184 Vail, CO 26-Mar-07 TISH SM139 Fort Collins, CO No Date TINC SM185 Vail, CO 26-Mar-07 TISH SM140 Loveland, CO No Date TISH SM186 Vail, CO 26-Mar-07 TISH SM141 Fort Collins, CO No Date TISH SM187 Vail, CO 26-Mar-07 TISH SM142 Fort Collins, CO No Date TISH SM188 Vail, CO 26-Mar-07 TISH SM143 Loveland, CO No Date TISH SM189 Vail, CO 26-Mar-07 TISH SM144 Loveland, CO No Date TISH SM190 Vail, CO 26-Mar-07 TISH SM145 Loveland, CO No Date TISH SM191 Vail, CO 26-Mar-07 TISH SM146 Loveland, CO No Date TISH SM192 Vail, CO 26-Mar-07 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM193 Vail, CO 26-Mar-07 TISH SM241 Vail, CO 26-Mar-07 TISH SM194 Vail, CO 26-Mar-07 TISH SM242 Vail, CO 26-Mar-07 TISH SM195 Vail, CO 26-Mar-07 TISH SM243 Vail, CO 26-Mar-07 TISH SM196 Vail, CO 26-Mar-07 TISH SM244 Vail, CO 26-Mar-07 TISH SM197 Vail, CO 26-Mar-07 TISH SM245 Vail, CO 26-Mar-07 TISH SM198 Vail, CO 26-Mar-07 TISH SM246 Vail, CO 26-Mar-07 TISH SM199 Vail, CO 26-Mar-07 TISH SM247 Vail, CO 26-Mar-07 TISH SM200 Vail, CO 26-Mar-07 TISH SM248 Vail, CO 26-Mar-07 TISH SM201 Vail, CO 26-Mar-07 TISH SM249 Vail, CO 26-Mar-07 TISH SM202 Vail, CO 26-Mar-07 TISH SM250 Vail, CO 26-Mar-07 TISH SM203 Vail, CO 26-Mar-07 TISH SM251 Vail, CO 26-Mar-07 TISH SM204 Vail, CO 26-Mar-07 TISH SM252 Vail, CO 26-Mar-07 TISH SM205 Vail, CO 26-Mar-07 TISH SM253 Vail, CO 26-Mar-07 TISH SM206 Vail, CO 26-Mar-07 TISH SM254 Vail, CO 26-Mar-07 TISH SM207 Vail, CO 26-Mar-07 TISH SM255 Vail, CO 26-Mar-07 TISH SM208 Vail, CO 26-Mar-07 TISH SM256 Vail, CO 26-Mar-07 TISH SM209 Vail, CO 26-Mar-07 TISH SM257 Vail, CO 26-Mar-07 TISH SM210 Vail, CO 26-Mar-07 TISH SM258 Vail, CO 26-Mar-07 TISH SM211 Vail, CO 26-Mar-07 TISH SM259 Vail, CO 26-Mar-07 TISH SM212 Vail, CO 26-Mar-07 TISH SM260 Vail, CO 26-Mar-07 TISH SM213 Vail, CO 26-Mar-07 TISH SM261 Vail, CO 26-Mar-07 TISH SM214 Vail, CO 26-Mar-07 TISH SM262 Vail, CO 26-Mar-07 TISH SM215 Vail, CO 26-Mar-07 TISH SM263 Vail, CO 26-Mar-07 TISH SM216 Vail, CO 26-Mar-07 TISH SM264 Vail, CO 26-Mar-07 TISH SM217 Vail, CO 26-Mar-07 TISH SM265 Vail, CO 26-Mar-07 TISH SM218 Vail, CO 26-Mar-07 TISH SM266 Vail, CO 26-Mar-07 TISH SM219 Vail, CO 26-Mar-07 TISH SM267 Vail, CO 26-Mar-07 TISH SM220 Vail, CO 26-Mar-07 TISH SM268 Vail, CO 26-Mar-07 TISH SM221 Vail, CO 26-Mar-07 TISH SM269 Vail, CO 26-Mar-07 TISH SM222 Vail, CO 26-Mar-07 TISH SM270 Vail, CO 26-Mar-07 TISH SM223 Vail, CO 26-Mar-07 TISH SM271 Vail, CO 26-Mar-07 TISH SM224 Vail, CO 26-Mar-07 TISH SM272 Vail, CO 26-Mar-07 TISH SM225 Vail, CO 26-Mar-07 TISH SM273 Vail, CO 26-Mar-07 TISH SM226 Vail, CO 26-Mar-07 TISH SM274 Vail, CO 26-Mar-07 TISH SM227 Vail, CO 26-Mar-07 TISH SM275 Vail, CO 26-Mar-07 TISH SM228 Vail, CO 26-Mar-07 TISH SM276 Vail, CO 26-Mar-07 TISH SM229 Vail, CO 26-Mar-07 TISH SM277 Vail, CO 26-Mar-07 TISH SM230 Vail, CO 26-Mar-07 TISH SM278 Vail, CO 26-Mar-07 TISH SM231 Vail, CO 26-Mar-07 TISH SM279 Vail, CO 26-Mar-07 TISH SM232 Vail, CO 26-Mar-07 TISH SM280 Vail, CO 26-Mar-07 TISH SM233 Vail, CO 26-Mar-07 TISH SM281 Vail, CO 26-Mar-07 TISH SM234 Vail, CO 26-Mar-07 TISH SM282 Vail, CO 26-Mar-07 TISH SM235 Vail, CO 26-Mar-07 TISH SM283 Vail, CO 26-Mar-07 TISH SM236 Vail, CO 26-Mar-07 TISH SM284 Vail, CO 26-Mar-07 TISH SM237 Vail, CO 26-Mar-07 TISH SM285 Vail, CO 26-Mar-07 TISH SM238 Vail, CO 26-Mar-07 TISH SM286 Vail, CO 26-Mar-07 TISH SM239 Vail, CO 26-Mar-07 TISH SM287 Vail, CO 26-Mar-07 TISH SM240 Vail, CO 26-Mar-07 TISH SM288 Vail, CO 26-Mar-07 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM289 Vail, CO 26-Mar-07 TISH SM336 Vail, CO 26-Mar-07 TISH SM290 Vail, CO 26-Mar-07 TISH SM337 Vail, CO 26-Mar-07 TISH SM291 Vail, CO 26-Mar-07 TISH SM338 Vail, CO 26-Mar-07 TISH SM292 Vail, CO 26-Mar-07 TISH SM339 Vail, CO 26-Mar-07 TISH SM293 Vail, CO 26-Mar-07 TISH SM340 Vail, CO 26-Mar-07 TISH SM294 Vail, CO 26-Mar-07 TISH SM341 Vail, CO 26-Mar-07 TISH SM295 Vail, CO 26-Mar-07 TISH SM342 Vail, CO 26-Mar-07 TISH SM296 Vail, CO 26-Mar-07 TISH SM343 Vail, CO 26-Mar-07 TISH SM297 Vail, CO 26-Mar-07 TISH SM344 Vail, CO 26-Mar-07 TISH SM298 Vail, CO 26-Mar-07 TISH SM345 Vail, CO 26-Mar-07 TISH SM299 Vail, CO 26-Mar-07 TISH SM346 Vail, CO 26-Mar-07 TISH SM300 Vail, CO 26-Mar-07 TISH SM347 Vail, CO 26-Mar-07 TISH SM301 Vail, CO 26-Mar-07 TISH SM348 Vail, CO 26-Mar-07 TISH SM302 Vail, CO 26-Mar-07 TISH SM349 Vail, CO 26-Mar-07 TISH SM303 Vail, CO 26-Mar-07 TISH SM350 Vail, CO 26-Mar-07 TISH SM304 Vail, CO 26-Mar-07 TISH SM351 Vail, CO 26-Mar-07 MNIV SM305 Vail, CO 26-Mar-07 TISH SM352 Vail, CO 26-Mar-07 TISH SM306 Vail, CO 26-Mar-07 TISH SM353 Vail, CO 26-Mar-07 TISH SM307 Vail, CO 26-Mar-07 TISH SM354 Vail, CO 26-Mar-07 TISH SM308 Vail, CO 26-Mar-07 TISH SM355 Vail, CO 26-Mar-07 TISH SM309 Vail, CO 26-Mar-07 TISH SM356 Vail, CO 26-Mar-07 TISH SM310 Vail, CO 26-Mar-07 TISH SM357 Vail, CO 26-Mar-07 TISH SM311 Vail, CO 26-Mar-07 TISH SM358 Vail, CO 26-Mar-07 TISH SM312 Vail, CO 26-Mar-07 TISH SM359 Vail, CO 26-Mar-07 TISH SM313 Vail, CO 26-Mar-07 TISH SM360 Vail, CO 26-Mar-07 TISH SM314 Vail, CO 26-Mar-07 TISH SM361 Vail, CO 26-Mar-07 TISH SM315 Vail, CO 26-Mar-07 TISH SM362 Vail, CO 26-Mar-07 TISH SM316 Vail, CO 26-Mar-07 TISH SM363 Vail, CO 26-Mar-07 TISH SM317 Vail, CO 26-Mar-07 TISH SM364 Vail, CO 26-Mar-07 TISH SM318 Vail, CO 26-Mar-07 TISH SM365 Vail, CO 26-Mar-07 TISH SM319 Vail, CO 26-Mar-07 TISH SM366 Vail, CO 26-Mar-07 TISH SM320 Vail, CO 26-Mar-07 TISH SM367 Vail, CO 26-Mar-07 TISH SM321 Vail, CO 26-Mar-07 TISH SM368 Vail, CO 26-Mar-07 TISH SM322 Vail, CO 26-Mar-07 TISH SM369 Breckenridge, CO 1-May-07 TISH SM323 Vail, CO 26-Mar-07 TISH SM370 Breckenridge, CO 1-May-07 TISH SM324 Vail, CO 26-Mar-07 TISH SM371 Breckenridge, CO 1-May-07 TISH SM325 Vail, CO 26-Mar-07 TISH SM372 Breckenridge, CO 1-May-07 TISH SM326 Vail, CO 26-Mar-07 TISH SM373 Breckenridge, CO 1-May-07 TISH SM327 Vail, CO 26-Mar-07 TISH SM374 Breckenridge, CO 1-May-07 TISH SM328 Vail, CO 26-Mar-07 TISH SM375 Breckenridge, CO 1-May-07 TISH SM329 Vail, CO 26-Mar-07 TISH SM376 Breckenridge, CO 1-May-07 TISH SM330 Vail, CO 26-Mar-07 TISH SM377 Breckenridge, CO 1-May-07 TISH SM331 Vail, CO 26-Mar-07 TISH SM378 Breckenridge, CO 1-May-07 TISH SM332 Vail, CO 26-Mar-07 TISH SM379 Breckenridge, CO 1-May-07 TISH SM333 Vail, CO 26-Mar-07 TISH SM380 Breckenridge, CO 1-May-07 TISH SM334 Vail, CO 26-Mar-07 TISH SM381 Breckenridge, CO 1-May-07 TISH SM335 Vail, CO 26-Mar-07 TISH SM382 Breckenridge, CO 1-May-07 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM383 Breckenridge, CO 1-May-07 TISH SM430 Breckenridge, CO 1-May-07 TISH SM384 Breckenridge, CO 1-May-07 TISH SM430 Breckenridge, CO 1-May-07 TISH SM385 Breckenridge, CO 1-May-07 TISH SM431 Breckenridge, CO 1-May-07 TISH SM386 Breckenridge, CO 1-May-07 TISH SM432 Breckenridge, CO 1-May-07 TISH SM387 Breckenridge, CO 1-May-07 TISH SM433 Breckenridge, CO 1-May-07 TISH SM388 Breckenridge, CO 1-May-07 TISH SM434 Breckenridge, CO 1-May-07 TISH SM389 Breckenridge, CO 1-May-07 TISH SM435 Breckenridge, CO 1-May-07 TISH SM390 Breckenridge, CO 1-May-07 TISH SM436 Breckenridge, CO 1-May-07 TISH SM391 Breckenridge, CO 1-May-07 TISH SM437 Breckenridge, CO 1-May-07 TISH SM392 Breckenridge, CO 1-May-07 TISH SM438 Breckenridge, CO 1-May-07 TISH SM393 Breckenridge, CO 1-May-07 TISH SM439 Breckenridge, CO 1-May-07 TISH SM394 Breckenridge, CO 1-May-07 TISH SM440 Breckenridge, CO 1-May-07 TISH SM395 Breckenridge, CO 1-May-07 TISH SM441 Breckenridge, CO 1-May-07 TISH SM396 Breckenridge, CO 1-May-07 TISH SM442 Breckenridge, CO 1-May-07 TISH SM397 Breckenridge, CO 1-May-07 TISH SM443 Breckenridge, CO 1-May-07 TISH SM398 Breckenridge, CO 1-May-07 TISH SM444 Breckenridge, CO 1-May-07 TISH SM399 Breckenridge, CO 1-May-07 TISH SM445 Breckenridge, CO 1-May-07 TISH SM400 Breckenridge, CO 1-May-07 TISH SM446 Breckenridge, CO 1-May-07 TISH SM401 Breckenridge, CO 1-May-07 TISH SM447 Breckenridge, CO 1-May-07 TISH SM402 Breckenridge, CO 1-May-07 TISH SM448 Breckenridge, CO 1-May-07 TISH SM403 Breckenridge, CO 1-May-07 TISH SM449 Breckenridge, CO 1-May-07 TISH SM404 Breckenridge, CO 1-May-07 TISH SM450 Breckenridge, CO 1-May-07 TISH SM405 Breckenridge, CO 1-May-07 TISH SM451 Breckenridge, CO 1-May-07 TISH SM406 Breckenridge, CO 1-May-07 TISH SM452 Breckenridge, CO 1-May-07 TISH SM407 Breckenridge, CO 1-May-07 TISH SM453 Breckenridge, CO 1-May-07 TISH SM408 Breckenridge, CO 1-May-07 TISH SM454 Breckenridge, CO 1-May-07 TISH SM409 Breckenridge, CO 1-May-07 TISH SM455 Breckenridge, CO 1-May-07 TISH SM410 Breckenridge, CO 1-May-07 TISH SM456 Breckenridge, CO 1-May-07 TISH SM411 Breckenridge, CO 1-May-07 TISH SM457 Breckenridge, CO 1-May-07 TISH SM412 Breckenridge, CO 1-May-07 TISH SM458 Breckenridge, CO 1-May-07 TISH SM413 Breckenridge, CO 1-May-07 TISH SM459 Breckenridge, CO 1-May-07 TISH SM414 Breckenridge, CO 1-May-07 TISH SM460 Breckenridge, CO 1-May-07 TISH SM415 Breckenridge, CO 1-May-07 TISH SM461 Breckenridge, CO 1-May-07 TISH SM416 Breckenridge, CO 1-May-07 TISH SM462 Breckenridge, CO 1-May-07 TISH SM417 Breckenridge, CO 1-May-07 TISH SM463 Breckenridge, CO 1-May-07 TISH SM418 Breckenridge, CO 1-May-07 TISH SM464 Breckenridge, CO 1-May-07 TISH SM419 Breckenridge, CO 1-May-07 TISH SM465 Breckenridge, CO 1-May-07 TISH SM420 Breckenridge, CO 1-May-07 TISH SM466 Breckenridge, CO 1-May-07 TISH SM421 Breckenridge, CO 1-May-07 TISH SM467 Breckenridge, CO 1-May-07 TISH SM422 Breckenridge, CO 1-May-07 TISH SM468 Breckenridge, CO 1-May-07 TISH SM423 Breckenridge, CO 1-May-07 TISH SM469 Breckenridge, CO 1-May-07 TISH SM424 Breckenridge, CO 1-May-07 TISH SM470 Breckenridge, CO 1-May-07 TISH SM425 Breckenridge, CO 1-May-07 TISH SM471 Breckenridge, CO 1-May-07 TISH SM426 Breckenridge, CO 1-May-07 TISH SM472 Breckenridge, CO 1-May-07 TISH SM427 Breckenridge, CO 1-May-07 TISH SM473 Breckenridge, CO 1-May-07 TISH SM428 Breckenridge, CO 1-May-07 TISH SM474 Breckenridge, CO 1-May-07 TISH SM429 Breckenridge, CO 1-May-07 TISH SM475 Breckenridge, CO 1-May-07 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM476 Breckenridge, CO 1-May-07 TISH SM523 Breckenridge, CO 1-May-07 TISH SM477 Breckenridge, CO 1-May-07 TISH SM524 Breckenridge, CO 1-May-07 TISH SM478 Breckenridge, CO 1-May-07 TISH SM525 Breckenridge, CO 1-May-07 TISH SM479 Breckenridge, CO 1-May-07 TISH SM526 Breckenridge, CO 1-May-07 TISH SM480 Breckenridge, CO 1-May-07 TISH SM527 Breckenridge, CO 1-May-07 TISH SM481 Breckenridge, CO 1-May-07 TISH SM528 Breckenridge, CO 1-May-07 TISH SM482 Breckenridge, CO 1-May-07 TISH SM529 Breckenridge, CO 1-May-07 TISH SM483 Breckenridge, CO 1-May-07 TISH SM530 Breckenridge, CO 1-May-07 TISH SM484 Breckenridge, CO 1-May-07 TISH SM531 Breckenridge, CO 1-May-07 TISH SM485 Breckenridge, CO 1-May-07 TISH SM532 Breckenridge, CO 1-May-07 TISH SM486 Breckenridge, CO 1-May-07 TISH SM533 Vail, CO 26-Mar-07 TISH SM487 Breckenridge, CO 1-May-07 TISH SM534 Breckenridge, CO 1-May-07 TISH SM488 Breckenridge, CO 1-May-07 TISH SM535 Breckenridge, CO 1-May-07 TISH SM489 Breckenridge, CO 1-May-07 TISH SM536 Vail, CO 26-Mar-07 TISH SM490 Breckenridge, CO 1-May-07 TISH SM537 Breckenridge, CO 1-May-07 TISH SM491 Breckenridge, CO 1-May-07 TISH SM538 Breckenridge, CO 1-May-07 TISH SM492 Breckenridge, CO 1-May-07 TISH SM539 Breckenridge, CO 1-May-07 TISH SM493 Breckenridge, CO 1-May-07 TISH SM540 Breckenridge, CO 1-May-07 TISH SM494 Breckenridge, CO 1-May-07 TISH SM541 Breckenridge, CO 1-May-07 TISH SM495 Breckenridge, CO 1-May-07 TISH SM542 Breckenridge, CO 1-May-07 TISH SM496 Breckenridge, CO 1-May-07 TISH SM543 Breckenridge, CO 1-May-07 TISH SM497 Breckenridge, CO 1-May-07 TISH SM544 Breckenridge, CO 1-May-07 TISH SM498 Breckenridge, CO 1-May-07 TISH SM545 Breckenridge, CO 1-May-07 TISH SM499 Breckenridge, CO 1-May-07 TISH SM546 Breckenridge, CO 1-May-07 TISH SM500 Breckenridge, CO 1-May-07 TISH SM547 Breckenridge, CO 1-May-07 TISH SM501 Breckenridge, CO 1-May-07 TISH SM548 Breckenridge, CO 1-May-07 TISH SM502 Breckenridge, CO 1-May-07 TISH SM549 Breckenridge, CO 1-May-07 TISH SM503 Breckenridge, CO 1-May-07 TISH SM550 Breckenridge, CO 1-May-07 TISH SM504 Breckenridge, CO 1-May-07 TISH SM551 Breckenridge, CO 1-May-07 TISH SM505 Breckenridge, CO 1-May-07 TISH SM552 Breckenridge, CO 1-May-07 TISH SM506 Breckenridge, CO 1-May-07 TISH SM553 Breckenridge, CO 1-May-07 TISH SM507 Vail, CO 26-Mar-07 TISH SM554 Breckenridge, CO 1-May-07 TISH SM508 Vail, CO 26-Mar-07 TISH SM555 Breckenridge, CO 1-May-07 TISH SM509 Vail, CO 26-Mar-07 TISH SM556 Breckenridge, CO 1-May-07 TISH SM510 Vail, CO 26-Mar-07 TISH SM557 Breckenridge, CO 1-May-07 TISH SM511 Vail, CO 26-Mar-07 TISH SM558 Breckenridge, CO 1-May-07 TISH SM512 Vail, CO 26-Mar-07 TISH SM559 Breckenridge, CO 1-May-07 TISH SM513 Breckenridge, CO 1-May-07 TISH SM560 Breckenridge, CO 26-Mar-07 TISH SM514 Breckenridge, CO 1-May-07 TISH SM561 Breckenridge, CO 1-May-07 TISH SM515 Breckenridge, CO 1-May-07 TISH SM562 Vail, CO 26-Mar-07 TISH SM516 Breckenridge, CO 1-May-07 TISH SM563 Breckenridge, CO 1-May-07 TISH SM517 Breckenridge, CO 1-May-07 TISH SM564 Breckenridge, CO 1-May-07 TISH SM518 Breckenridge, CO 1-May-07 TISH SM565 Breckenridge, CO 1-May-07 TISH SM519 Breckenridge, CO 1-May-07 TISH SM566 Breckenridge, CO 1-May-07 TISH SM520 Breckenridge, CO 1-May-07 TISH SM567 Breckenridge, CO 1-May-07 TISH SM521 Breckenridge, CO 1-May-07 TISH SM568 Breckenridge, CO 1-May-07 TISH SM522 Breckenridge, CO 1-May-07 TISH SM569 Breckenridge, CO 1-May-07 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM570 Breckenridge, CO 1-May-07 TISH SM617 Canada Acquired 2007 TISH SM571 Breckenridge, CO 1-May-07 TISH SM618 Canada Acquired 2007 TINC SM572 Breckenridge, CO 1-May-07 TISH SM619 Canada Acquired 2007 TINC SM573 Breckenridge, CO 1-May-07 TISH SM620 Canada Acquired 2007 TISH SM574 Breckenridge, CO 1-May-07 TISH SM621 Canada Acquired 2007 TISH SM575 Breckenridge, CO 1-May-07 TISH SM622 Canada Acquired 2007 TISH SM576 Breckenridge, CO 1-May-07 TISH SM623 Canada Acquired 2007 TISH SM577 Breckenridge, CO 1-May-07 TISH SM624 Breckenridge, CO 1-May-07 TISH SM578 Breckenridge, CO 1-May-07 TISH SM625 Breckenridge, CO 1-May-07 TISH SM579 Breckenridge, CO 1-May-07 TISH SM626 Breckenridge, CO 1-May-07 TISH SM580 Breckenridge, CO 1-May-07 TISH SM627 Vail, CO 26-Mar-07 TISH SM581 Vail, CO 26-Mar-07 TINC SM628 Edwards, CO 12-14 May 2007 TINC SM582 Vail, CO 26-Mar-07 TISH SM629 Wolcott, CO 12-14 May 2007 MBOR SM583 Vail, CO 26-Mar-07 TINC SM630 Edwards, CO 12-14 May 2007 TPHA SM584 Vail, CO 26-Mar-07 TINC SM631 Telluride, CO 12-14 May 2007 TISH SM585 Vail, CO 26-Mar-07 TINC SM632 Basalt, CO 12-14 May 2007 TISH SM587 Vail, CO 26-Mar-07 TINC SM633 Telluride, CO 12-14 May 2007 TINC SM588 Vail, CO 26-Mar-07 TINC SM634 Wolcott, CO 12-14 May 2007 TINC SM589 Vail, CO 26-Mar-07 TINC SM635 Wolcott, CO 12-14 May 2007 TINC SM590 Vail, CO 26-Mar-07 TINC SM636 Wolcott, CO 12-14 May 2007 TINC SM591 Vail, CO 26-Mar-07 TINC SM637 Telluride, CO 12-14 May 2007 TINC SM592 Vail, CO 26-Mar-07 TINC SM638 Telluride, CO 12-14 May 2007 TINC SM593 Michigan Acquired 2007 TISH SM639 Edwards, CO 12-14 May 2007 TINC SM594 Wisconsin Acquired 2007 TISH SM640 Granby, CO 12-14 May 2007 TINC SM595 Michigan Acquired 2007 TISH SM641 Vail, CO 12-14 May 2007 TISH SM596 Michigan Acquired 2007 TISH SM642 Vail, CO 12-14 May 2007 TISH SM597 Wisconsin Acquired 2007 TISH SM643 Vail, CO 12-14 May 2007 TISH SM598 Wisconsin Acquired 2007 TISH SM644 Vail, CO 12-14 May 2007 TISH SM599 Wisconsin Acquired 2007 TISH SM645 Granby, CO 12-14 May 2007 WCIR SM600 Michigan Acquired 2007 TINC SM646 Telluride, CO 12-14 May 2007 TISH SM601 Minnesota Acquired 2007 TINC SM647 Basalt, CO 12-14 May 2007 TISH SM602 Wisconsin Acquired 2007 TINC SM648 Edwards, CO 12-14 May 2007 TISH SM603 Michigan Acquired 2007 TINC SM649 Vail, CO 12-14 May 2007 TISH SM604 Michigan Acquired 2007 TISH SM650 Vail, CO 12-14 May 2007 TISH SM605 Michigan Acquired 2007 TISH SM651 Vail, CO 12-14 May 2007 TISH SM606 Michigan Acquired 2007 TISH SM652 Vail, CO 12-14 May 2007 TISH SM607 Idaho Acquired 2007 TINC SM653 Vail, CO 12-14 May 2007 TISH SM608 Idaho Acquired 2007 TINC SM654 Vail, CO 12-14 May 2007 TISH SM609 Idaho Acquired 2007 TINC SM655 Vail, CO 12-14 May 2007 TISH SM610 Idaho Acquired 2007 TINC SM656 Vail, CO 12-14 May 2007 TISH SM611 Idaho Acquired 2007 TISH SM657 Vail, CO 12-14 May 2007 TISH SM612 Idaho Acquired 2007 TISH SM658 Edwards, CO 12-14 May 2007 TPHA SM613 Idaho Acquired 2007 TISH SM659 Edwards, CO 12-14 May 2007 TISH SM614 Idaho Acquired 2007 TISH SM660 Edwards, CO 12-14 May 2007 TISH SM615 Canada Acquired 2007 TISH SM661 Basalt, CO 12-14 May 2007 TISH SM616 Canada Acquired 2007 TISH SM662 Telluride, CO 12-14 May 2007 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

Isolate No. Collected From Date Collected Idx Isolate No. Collected From Date Collected Idx SM663 Edwards, CO 12-14 May 2007 TISH SM664 Keystone, CO 12-14 May 2007 TISH SM665 Wolcott, CO 12-14 May 2007 TISH SM667 Granby, CO 12-14 May 2007 TISH SM668 Granby, CO 12-14 May 2007 TISH SM669 Edwards, CO 12-14 May 2007 TPHA SM670 Edwards, CO 12-14 May 2007 TISH SM671 Granby, CO 12-14 May 2007 TISH SM672 Edwards, CO 12-14 May 2007 TISH SM673 Vail, CO 12-14 May 2007 TISH SM674 Granby, CO 12-14 May 2007 TISH SM675 Granby, CO 12-14 May 2007 TISH SM676 Keystone, CO 12-14 May 2007 TISH SM677 Vail, CO 12-14 May 2007 TISH SM678 Vail, CO 12-14 May 2007 TISH SM679 Vail, CO 12-14 May 2007 TISH SM680 Vail, CO 12-14 May 2007 TISH SM681 Telluride, CO 12-14 May 2007 TISH SM682 Keystone, CO 12-14 May 2007 TISH SM683 Keystone, CO 12-14 May 2007 TISH X Identification of Typhula isolates; TISH = Typhula ishikariensis, TINC = Typhula incarnata, TPHAC = Typhula phacorrhiza, MNIV = Microdochium nivale, MBOR = Myriosclerotinia borealis, and WCIR = Waitea circinata.

APPENDIX 2

FUNGICIDE TRIAL RESULTS FOR BRECKENRIDGE, CO 2004-05 AND 2005-06

FUNGICIDE TRIAL RESULTS FOR VAIL, CO 2004-05, 2005-06, 2006-07, 2007-08, 2008-09, 2009-10, 2010-11

KENTUCKY BLUEGRASS (Poa pratensis cv. unknown) T.D. Blunt, N.A. Tisserat and S. Fichtner ANNUAL BLUEGRASS (Poa annua cv. unknown) Bioagricultural Sciences and Pest Management Pink Snow Mold (Michrodium nivale) Colorado State University Gray Snow Mold (Typhula spp.) Fort Collins, CO 80523-1177

Fungicide evaluation for the preventive control of snow molds on Kentucky Bluegrass and Annual Bluegrass, 2004-2005.

Experiments for control of pink and gray snow mold were performed on a Kentucky Bluegrass/Annual Bluegrass fairway at Breckenridge Golf Club, Fairway 4 Beaver, Breckenridge, CO (average elevation 9600 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of three treatments of fungicides were applied 1 Oct 04. A second application of the three treatments and the remainder of the treatments were applied 18 Oct 04. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 04 May 05.

Evaluations of the plots were later than the Vail plots due to the fact the amount of snow cover received in the central Rocky Mountains was substantial. The plots were evaluated differently this year than in previous years. Our rating system was change to a percent of disease incidence per treatment that more accurately represents the actual disease present in each treatment/plot. The plots at Breckenridge, CO showed results consistent with disease pressure but not as severe of disease pressure as Vail. The Breckenridge course is mainly Kentucky Bluegrass which differs from Vail (mainly Poa annua) Results were calculated using SAS system proc glm calculating the means of the four replications and using Fisher’s Protected Least Significant Difference (α=0.05).

Table 1. Snow mold fungicide trials at Breckenridge Golf Club, Breckenridge, CO______Treatment and Rate/1000 sq. ft. Disease Rating_ CL-EXP-2 4.0 oz + Daconil WS 6FL 5.5 fl oz ...... 1.38k CL-EXP-4 1.0 oz + Spectro 90WDG 5.76 oz ...... 1.88jk Syngenta Eperimental A14036 43.3SE 9.3 fl oz ...... 2.00jk Spectro 90WDG 4.0 oz FB Spectro 90WDG 4.0 oz + CL-EXP-4 1.0 oz ...... 2.50i-k Medallion 50WDG 0.142 oz + Daconil WS 6FL 4.80 fl oz + BannerMAXX 1.3MEC 1.71 fl oz ...... 3.38 i-k Syngenta Experimental A14036 43.3SE 18.6 fl oz...... 4.50h-k CL-EXP-2 4.0 oz + Daconil WS 6FL 5.5 fl oz + Magnum 3.5 fl oz ...... 4.50h-k Chipco 26GT 4.0 fl oz + Signature 80WDG 8.0 oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 5.25g-k Endorse 2.5WP 4.0 oz + Spectro 90WDG 5.76 oz ...... 5.63f-k Syngenta Experimental A14212C 1.67EC 5.24 fl oz ...... 6.63f-k BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz ...... 6.75f-k Syngenta Experimental A14036 43.3SE 4.7 fl oz...... 7.38f-k Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz + Turfcide 400 8.0 fl oz ...... 7.50e-k Daconil WS 6FL 5.5 fl oz + Medallion 50WDG 0.5 oz ...... 7.50e-k BannerMAXX 1.3MEC 4.0 fl oz FB BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz .... 8.63e-k Chipco 26GT 4.0 fl oz + Compass 50WDG 0.25 oz + Signature 80WDG 8.0 oz + Turfcide 400 8.0 fl oz ...... 10.13e-k Syngenta Experimental A14212C 1.67EC 3.0 fl oz ...... 10.63e-k AND 4322G 10.0 lb ...... 11.25e-k Chipco 26GT 4.0 fl oz + Signature 80WDG 8.0 oz + Turfcide 400 8.0 fl oz ...... 12.63e-k

AND 3118G 6.0 lb + AND 3237G 12.0 lb + AND 3238G 7.2 lb ...... 12.75e-k Spectro 90WDG 4.0 oz FB Spectro 90WDG 4.0 oz + Endorse 2.5WP 4.0 oz ...... 15.25d-j AND 4311G 6.6 lb ...... 16.25d-i AND 4309G 6.0 lb + AND 3237G 12.0 lb + AND 3238G 7.2 lb ...... 17.50d-h AND 4323G 6.6 lb ...... 18.63c-g Heritage 50WDG 0.4 oz + Medallion 50WDG 0.5 oz ...... 19.38c-f AND 4333G 9.0 lb ...... 19.38c-f Chipco 26GT 4 fl oz + Compass 50WDG 0.25 oz + Turfcide 400 8.0 fl oz ...... 19.38c-f Eagle 1.67EW 2.4 oz ...... 21.25b-e AND 4310G 8.0 lb ...... 27.50b-d AND 4334G 9.0 lb ...... 28.75b-d AND3223G 6.6 lb ...... 31.88bc Eagle 1.67EW 1.4 oz ...... 33.88b Untreated Control ...... 61.88a LSD (α=0.05) ...... 13.78_

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ANNUAL BLUEGRASS (Poa annua cv. unknown) T.D. Blunt, N. A. Tisserat and S. Fichtner Pink Snow Mold (Michrodium nivale) Bioagricultural Sciences and Pest Management Gray Snow Mold (Typhula spp.) Colorado State University Fort Collins, CO 80523-1177

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2004-2005.

Experiments for control of pink and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of three treatments of fungicides were applied 1 Oct 04. A second application of the three treatments and the remainder of the treatments were applied 15 Oct 04. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 22 Apr 05.

Evaluations of the plots were about average for Vail even though the amount of snow cover received in the central Rocky Mountains was substantial. The plots were evaluated differently this year than in previous years. Our rating system was change to a percent of disease incidence per treatment that more accurately represents the actual disease present in each treatment/plot. The plots at Vail, CO showed results consistent with increased disease pressure as evidenced by the untreated control results. Results were calculated using SAS system proc glm calculating the means of the four replications and using Fisher’s Protected Least Significant Difference (α=0.05).

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. Disease Rating_ Chipco 26GT 4 fl oz + Daconil Ultrex 82.5WDG 3.2 oz + Turfcide 400 8 fl oz ...... 2.38o Spectro 90WDG 4.0 oz FB Spectro 90WDG 4.0 oz + CL-EXP-4 1.0 oz ...... 3.88o Spectro 90WDG 4.0 oz FB Spectro 90WDG 4.0 oz + Endorse 2.5WP 4.0 oz ...... 5.75no Spectro 90WDG 5.76 oz + CL-EXP-4 1.0 oz ...... 6.13no Daconil WS 6FL 5.5 fl oz + CL-EXP-2 4.0 oz + Magnum 3.5 fl oz...... 6.88m-o Daconil WS 6FL 5.5 fl oz + CL-EXP-2 4.0 oz ...... 9.00l-o Chipco 26GT 4.0 fl oz + Compass 50WDG 0.25 oz + Signature 80WDG 8.0 oz + Turfcide 400 8.0 fl oz ...... 10.00l-o Spectro 90WDG 5.76 oz + Endorse 2.5WP 4.0 oz ...... 10.00l-o Daconil WS 6FL 5.5 fl oz + Medallion 50WDG 0.5 oz ...... 11.25k-o Chipco 26GT 4.0 fl oz + Compass 50WDG 0.25 oz + Turfcide 400 8.0 fl oz...... 13.75k-o Syngenta Experimental A14036 43.3SE 18.7 fl oz...... 18.13k-o Syngenta Experimental A14036 43.3SE 9.3 fl oz...... 19.38j-o Chipco 26GT 4.0 fl oz + Signature 80WDG 8.0 oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 20.63h-o Medallion 50WDG 0.142 oz + Daconil WS 6FL 4.8 fl oz + BannerMAXX 1.3MEC 1.71 fl oz ...... 22.50h-m BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz ...... 25.00g-m Chipco 26GT 4.0 fl oz + Signature 80WDG 8.0 oz + Turfcide 400 8.0 fl oz ...... 25.63f-l Syngenta Experimental A14036 43.3SE 4.7 fl oz...... 25.63f-l AND 3223G 6.6 lb ...... 25.63f-l BannerMAXX 1.3MEC 4.0 fl oz FB BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WDG 0.5 oz .... 28.375f-k AND 4311G 6.6 lb ...... 29.38f-k AND 4322G 10 lb ...... 32.50g-j Heritage 50WDG 0.4 oz + Medallion 50WDG 0.5 oz ...... 32.50g-j

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AND 4323G 6.6 lb ...... 36.88e-i Syngenta Experimental A14212C 167EC 5.24 fl oz ...... 38.13e-h AND 4309F 6.0 lb + AND 3237G 12.0 lb + AND 3238G 7.2 lb ...... 43.13e-g AND 3118G 6.0 lb + AND 3237G 12.0 lb + AND 3238G 7.2 lb ...... 43.75ef Syngenta Experimental A14212C 167EC 3.0 fl oz ...... 51.88de AND 4334G 9.0 lb ...... 62.50cd AND 4310G 8.0 lb ...... 65.63cd AND 4333G 9.0 lb ...... 70.63bc Eagle 1.67EW 2.4 fl oz ...... 86.25ab Eagle 1.67EW 1.4 fl oz ...... 89.38a Untreated Control ...... 93.75a LSD (α=0.05) ...... 18.37_

102

KENTUCKY BLUEGRASS (Poa pratensis cv. unknown) T.D. Blunt, N.A. Tisserat ANNUAL BLUEGRASS (Poa annua cv. unknown) Bioagricultural Sciences and Pest Management Pink Snow Mold (Michrodium nivale) Colorado State University Gray Snow Mold (Typhula spp.) Fort Collins, CO 80523-1177

Fungicide evaluation for the preventive control of snow molds on Kentucky Bluegrass, 2005-2006.

Experiments for control of pink and gray snow mold were performed on a Kentucky Bluegrass fairway at Breckenridge Golf Club, Fairway 4 Beaver, Breckenridge, CO (average elevation 9300 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of three treatments of fungicides were applied 9 Sept 05. A second application of the three treatments and the remainder of the treatments were applied 24 Oct 05. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 03 May 06.

Evaluations of the plots were about average for Breckenridge even though the amount of snow cover received in the central Rocky Mountains was substantial. The plots were evaluated this year based on a percent of disease incidence per treatment that more accurately represents the actual disease present in each treatment/plot. The plots at Breckenridge, CO showed results consistent with increased disease pressure as evidenced by the untreated control results, although Breckenridge had less overall disease incidence than Vail. There was also significant vole damage on the trial plots. Results were calculated using SAS system proc glm calculating the means of the four replications and using Fisher’s Protected Least Significant Difference (α=0.05).

Table 1. Snow mold fungicide trials at Breckenridge Golf Club, Breckenridge, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Instrata 3.61SE 11.0 fl oz ...... 0.00k Headway 167EC 5.25 fl oz + Daconil WeatherStik 6F 5.5 fl oz ...... 0.00k Insignia 20WP 0.7 oz + Rever 4F 8.0 fl oz + Manicure Ultrex 82.5WDG 3.2 oz...... 0.00k Medallion 50WP 0.14 oz + Daconil WeatherStik 6F 2.36 fl oz + Banner MAXX 1.3ME 1.7 fl oz ... 0.25k Insignia 20WP 0.7 oz + Iprodione Pro 2SE 4.0 fl oz + Manicure Ultrex 82.5WDG 3.2 oz ...... 0.25k Cleary 26/36 4.0 fl oz + Endorse 2.5WP 4.0 oz + Daconil Ultrex 82.5WDG 5.5 oz...... 0.25k Endorse 2.5WP 4.0 oz + Spectro 90WDG 5.75 oz ...... 0.50jk Cleary 26/36 4.0 fl oz + CL-EXP-4 1.0 oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 0.50jk BannerMAXX 1.3ME 2.0 fl oz + Daconil WeatherStik 6F 5.5 fl oz ...... 0.75jk Insignia 20WDG 0.9 oz + Iprodione Pro 2SE 4.0 fl oz + Manicure Ultrex 82.5WDG 3.2 oz ...... 0.75jk 18 Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz + Revere 4F 12.0 fl oz ...... 0.75jk Instrata 3.61SE 5.5 fl oz FB Instrata 3.61SE 5.5 fl oz ...... 1.00jk Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz + Turfcide 400 8.0 fl oz ...... 1.25jk Insignia 20WDG 0.7 oz FB 18 Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz ...... 1.25jk Spectator Ultra 1.3ME 4.0 fl oz + Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz ... 1.25jk CL-EXP-4 1.0 oz + Spectro 90WDG 5.75 oz ...... 1.25jk Insignia 20WDG 0.9 oz + Manicure Ultrex 82.5WDG 3.2 oz ...... 1.50jk Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 1.50jk Rubigan AS 8.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 2.75i-k Cleary 26/36 4.0 fl oz + Endorse 2.5WP 4.0 oz ...... 3.00h-k Instrata 3.61SE 9.3 fl oz ...... 3.75h-k

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Insignia 20WDG 0.9 oz + Iprodione Pro 2SE 4.0 fl oz + Revere 4F 8.0 fl oz ...... 3.75h-k Instrata 3.61SE 4.7 fl oz ...... 5.00g-k Heritage TL 0.8ME 2.0 fl oz + Medallion 50WP 0.5 oz + Daconil WeatherStik 6F 5.5 fl oz ...... 5.00g-k Daconil WeatherStik 6F 5.5 fl oz + Medallion 50WP 0.5 oz ...... 8.25f-k Turfcide 400 8.0 fl oz + Daconil Ultrex 82.5 WDG 3.2 oz ...... 8.75f-k 18 Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz FB Revere 4F 12.0 fl oz ...... 9.25f-k Manicure Ultra 82.5WDG 5.0 oz FB PCNB 12.5 Plus 10-3-23E 6 lb...... 9.25f-k Instrata 3.61SE 6.0 fl oz ...... 9.50f-k Chipco 26GT 4.0 fl oz + Lynx 22FL 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 9.50f-k Headway 167EC 5.25 fl oz ...... 10.00f-k Heritage TL 0.8ME 2.0 fl oz + Medallion 50WP 0.5 oz ...... 10.25f-k Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz + Revere 4F 12.0 fl oz ...... 10.50f-k Daconil Ultrex 82.5WDG 5.0 oz...... 10.75f-k Banner MAXX 1.3ME 4.0 fl oz FB Banner MAXX 1.3ME 4.0 fl oz + Medallion 50WP 0.5 oz ...... 11.25f-k Rubigan AS 8.0 fl oz + Heritage 50WDG 0.4 oz ...... 12.50f-k Turfcide 400 12.0 fl oz + Lysone 20 ml/100 ft2 ...... 12.50f-k Rubigan AS 8.0 fl oz + Turfcide 400 8.0 fl oz ...... 12.75f-k Turfcide 400 8.0 fl oz + Chipco 26GT 4.0 fl oz ...... 13.25f-k Revere 4F 12.0 fl oz ...... 14.00f-k Tartan 25SC 2.0 fl oz + Turfcide 400 8.0 fl oz ...... 14.50e-k Daconil Ultrex 82.5WDG 5.0 oz + Lysone 20 ml/100 ft2...... 14.50e-k Headway 167EC 5.25 fl oz + Medallion 50WP 0.5 oz ...... 15.75d-k Rubigan AS 8.0 fl oz ...... 16.50d-k Turfcide 400 16.0 fl oz ...... 17.50d-k Spectator Ultra 1.3ME 4.0 fl oz FB Revere 4F 12.0 fl oz ...... 17.75d-k Plant Helper 423 oz wt/a FB Plant Helper 423 oz wt/a ...... 20.75d-k Banner MAXX 1.3ME 2.0 fl oz + Turfcide 400 6.0 fl oz ...... 21.75b-k Turfcide 400 8.0 fl oz ...... 23.25b-k Turfcide 400 12.0 fl oz ...... 23.75b-j Turfcide 400 8.0 fl oz + Lysone 20 ml/100 ft2 ...... 25.00b-i Chipco 26GT 4.0 fl oz + Lynx 22FL 1.0 fl oz + Signature 80WDG 4.0 oz ...... 26.25b-h Banner MAXX 1.3ME 4.0 fl oz + Medallion 50WP 0.5 oz ...... 27.75b-g Chipco 26GT 4.0 fl oz + Lynx 22FL 1.0 fl oz ...... 29.75b-f Lysone 20 ml/100 ft2 ...... 31.25b-f Armada 50WP 1.2 oz FB Revere 4F 12.0 fl oz ...... 37.50b-e Untreated Control ...... 38.75b-d Plant Helper 1 lb a.i./100 gal FB Plant Helper 1 lb a.i./100 gal ...... 43.75a-c Untreated Control ...... 45.00ab Headway 167EC 3.0 fl oz FB Headway 167EC 3.0 fl oz ...... 64.00a LSD (α=0.05) ...... 23.37_

104

ANNUAL BLUEGRASS (Poa annua cv. unknown) T.D. Blunt and N. A. Tisserat Speckled snow mold (Typhula ishikariensis) Bioagricultural Sciences and Pest Management Gray Snow Mold (Typhula incarnata) Colorado State University Fort Collins, CO 80523-1177

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2005-2006.

Experiments for control of speckled and gray snow mold were performed on a Kentucky Bluegrass/Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of three treatments of fungicides were applied 9 Sept 05. A second application of the three treatments and the remainder of the treatments were applied 10 Oct 05. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 28 Apr 06.

Evaluations of the plots were about average for Vail even though the amount of snow cover received in the central Rocky Mountains was substantial. The plots were evaluated this year based on a percent of disease incidence per treatment that more accurately represents the actual disease present in each treatment/plot. The plots at Vail, CO showed results consistent with increased disease pressure as evidenced by the untreated control results. Results were calculated using SAS system proc glm calculating the means of the four replications and using Fisher’s Protected Least Significant Difference (α=0.05).

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ 18Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz + Revere 4F ...... 15.25v Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz + Turfcide 400 8.0 fl oz ...... 15.88uv Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz + Revere 4F 12.0 fl oz ...... 17.50uv BannerMAXX 1.3MEC 4.0 fl oz FB BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WP 0.5 oz ...... 19.63uv Insignia 20WDG 0.7 oz + Revere 4F 8.0 fl oz + Manicure Ultra 82.5WDG 3.2 oz ...... 21.38uv Instrata 3.61SE 5.5 fl oz FB Instrata 5.5 3.61SE 5.5 fl oz ...... 24.63t-v Cleary 26/36 4.0 fl oz + CL-EXP-4 1.0 oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 25.88s-v CL-EXP-4 1.0 oz + Spectro 90WDG 5.75 oz ...... 26.13 s-v Headway 167EC 5.25 fl oz + Medallion 50WP 0.5 oz ...... 27.00r-v Cleary 26/36 4.0 fl oz + Endorse 2.5WP 4.0 oz + Daconil Ultrex 82.5WDG 5.5 oz...... 29.25r-v Tartan 25SC 2.0 fl oz + Turfcide 400 8.0 fl oz ...... 31.75q-v BannerMAXX 1.3MEC 4.0 fl oz + Medallion 50WP 0.5 oz ...... 31.75q-v Insignia 20WDG 0.9 oz + Iprodione Pro 2SE 4.0 fl oz + Revere 4F 8.0 fl oz ...... 32.00p-v Insignia 20WDG 0.9 oz + Iprodione Pro 2SE 4.0 fl oz + Manicure Ultra 82.5WDG 3.2 oz ...... 32.50p-v Instrata 3.61SE 11.0 fl oz ...... 36.25o-v Headway 167EC 5.25 fl oz ...... 37.38n-v Insignia 20WP 0.7 oz + Iprodione Pro 2SE 4.0 fl oz + Manicure Ultra 82.5WDG 3.2 oz ...... 39.75m-v Chipco 26GT 4.0 fl oz + Lynx 22F 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 41.63l-u Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 50.00k-t Endorse 2.5WP 4.0 oz + Spectro 90WDG 5.75 oz ...... 50.38k-t Rubigan AS 8.0 fl oz + Heritage 50WDG 0.4 oz ...... 51.88j-g Revere 4000f 12.0 fl oz ...... 53.00k-r

105

Headway 167EC 5.25 fl oz + Daconil WS 6F 5.5 fl oz ...... 56.25i-q Instrata 3.61SE 9.3 fl oz ...... 56.88h-q Heritage TL 0.8ME 2.0 fl oz + Medallion 50WP 0.5 oz+ Daconil WeatherStik 6F 5.5 fl oz ...... 56.88h-q Turfcide 400 12.0 fl oz + Lysone 20ml/100ft2 ...... 57.75g-q Hertiage TL 0.8ME 2.0 fl oz + Medallion 50WP 0.5 oz ...... 57.75g-q Spectator Ultra 1.3MEC 4.0 fl oz + Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 fl oz 57.88g-q Turfcide 400 8.0 fl oz + Chipco 26GT 4.0 fl oz ...... 58.13g-p Instrata 3.61SE 6.0 fl oz ...... 58.75g-o Turfcide 400 12.0 fl oz ...... 58.75g-o Turfcide 400 16.0 fl oz ...... 59.25g-o Insignia 20WDG 0.7 oz FB 18Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz ...... 63.38f-n Daconil WeatherStik 6F 5.5 fl oz + Medallion 50WP 0.5 oz ...... 63.63f-m Armada 50WP 1.2 oz FB Revere 4F 12.0 fl oz ...... 63.75f-m Spectator Ultra 1.3MEC 4.0 fl oz FB Revere 4F 12.0 fl oz ...... 66.25e-k Rubigan AS 8.0 fl oz + Turfcide 400 8.0 fl oz ...... 66.63f-l Chipco 26GT 4.0 fl oz + Lynx 22FL 1.0 fl oz + Signature 80WDG 4.0 oz ...... 66.88f-l Turfcide 400 8.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 67.38d-l Cleary 26/36 4.0 fl oz + Endorse2.5WP 4.0 oz ...... 69.25c-k Untreated Control ...... 72.00b-k Insignia 20WDG 0.9 oz + Manicure Ultrex 82.5WDG 3.2 oz ...... 74.00a-k Headway 167EC 3.0 fl oz FB Headway 167EC 3.0 fl oz ...... 75.63a-k Turfcide 400 8.0 fl oz + Lysone 20 ml/100 ft2 ...... 77.13a-j Instrata 3.61SE 4.7 fl oz ...... 79.75a-i Untreated Control ...... 80.75a-i Turfcide 400 8.0 fl oz ...... 81.38a-i Medallion 50WP 0.14 oz + Daconil WeatherStik 6f 2.36 fl oz + Banner MAXX 1.3ME 1.7 fl oz ... 82.13a-i Banner MAXX 1.3ME 2.0 fl oz + Turfcide 400 6.0 fl oz ...... 82.13a-i Chipco 26GT 4.0 fl oz + Lynx 22FL 1.0 fl oz ...... 82.50a-h Banner MAXX 1.3ME 2.0 fl oz + Daconil WeatherStik 6F 5.5 fl oz ...... 83.75a-g 18Plus 23.3F 4.0 fl oz + Manicure Ultra 1.3ME 5.0 fl oz FB Revere 4F 12.0 fl oz ...... 83.75a-g Manicure Ultra 1.3ME 5.0 fl oz FB PCNB 12.5 Plus 10-3-23E 6 lb ...... 87.50a-f Rubigan AS 8.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 89.38a-f Plant Helper 1 lb a.i./100 gal FB Plant Helper 1 lb a.i./100 gal ...... 92.13a-e Rubigan AS 8.0 fl oz ...... 93.50a-d Daconil Ultrex 82.5WDG 5.0 oz...... 95.00a-c Lysone 20ml/100 ft2 ...... 95.63ab Plant Helper 423 oz wt/z FB Plant Helper 423 oz wt/a ...... 96.00ab Daconil Ultrex 82.5WDG 5.0 oz + Lysone 20 ml/100 ft2 ...... 98.75a LSD (α=0.05) ...... 25.85_

106

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2006-2007.

Experiments for control of speckled and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of three treatments of fungicides were applied 26 Sept 06. A second application of the three treatments and the remainder of the treatments were applied 24 Oct 06. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 26 Mar 07.

Snow had to be removed from plots prior to fungicide application in October. After application plots remained snow-free for two weeks. Snowmelt in late March occurred approximately two weeks earlier than normal. Nevertheless, disease pressure was hit with untreated plots averaging 89% plot area damaged. All fungicides except DPX-H6573 at 0.44 fl oz, DPX-LEM17 at 0.5 and 1.0 oz and DPX- YT669 at 0.425 fl oz reduced severity. However, most fungicide combinations did not provide acceptable levels of control (<10% damage). Plots were re-evaluated on 1 May 07 (results not shown) with most plots showing an increase in turf recovery from snow mold damage.

Evaluations of the plots were about average for Vail even though the amount of snow cover received in the central Rocky Mountains was substantial. The plots were evaluated based on a percent of disease incidence per treatment that more accurately represents the actual disease present in each treatment/plot. The plots at Vail, CO showed results consistent with increased disease pressure as evidenced by the untreated control results. Results were calculated using SAS system proc glm calculating the means of the four replications and using Fisher’s Protected Least Significant Difference (α=0.05).

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz + Revere 4F 12 fl oz ...... 6.63n Spectro 90WDG 4.0 oz FB Cleary 26/36 4 fl oz + Endorse 2.5WP 4.0 oz ...... 8.00nm Lynx 22FL 1.0 fl oz + Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5 WDG 5.0 oz ...... 8.63l-n Cleary 26/36 4.0 fl oz + CL-EXP-9 1.2 oz ...... 8.75l-n Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + CL-EXP-9 1.2 oz ...... 9.00l-n Insignia 20WDG 0.7 oz + Revere 4F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz ...... 9.00l-n Cleary 26/36 4.0 fl oz + Endorse 2.5 WP 4.0 WP ...... 9.50k-n Tartan 288SC 2.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 10.00k-n TBZ + TFS Green 2.0 fl oz + Chipco 26GT 4.0 fl oz ...... 14.38j-n Insignia 20WDG 0.7 oz + BAS595 1.0 oz + Manicure Ultrex 82.5WDG 5.0 oz ...... 15.50j-n Spectro 90WDG 5.75 oz + CL-EXP-9 1.2 oz ...... 16.50j-n Tartan 288SC 2.0 fl oz + Chipco 26GT 6.0 fl oz ...... 16.75j-n Instrata 3.61SE 11.0 fl oz ...... 18.00i-n TBZ + TFS Green 2.0 fl oz ...... 18.75i-n

107

Banner MAXX 1.3ME 2.0 fl oz + Daconil WeatherStik 6F 5.5 fl oz + SYNC ...... 19.63h-n DPX-LEM17 20SC 2.0 fl oz ...... 21.25h-n Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz + 18Plus 23.3F 4.0fl oz ...... 21.25h-n Tartan 288SC 2.0 fl oz + Prostar 70DF 2.2 fl oz ...... 22.75g-n Insignia 20WDG 0.7 oz + Revere 4F 4.0 fl oz + Manicure Ultra 82.5WDG 4.0 oz ...... 23.00g-n Instrata 3.61SE 9.0 fl oz ...... 23.50g-n Medallion 50WP 0.15 oz + Daconil WeatherStik 6f 2.5 fl oz + Banner Maxx 1.3ME 1.8fl oz ...... 25.00g-n Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 26.88f-n 18 Plus 23.3F 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz + Revere 4F 12.0 fl oz ...... 27.25f-n Banner MAXX 1.3ME 2.0 fl oz + Daconil WeatherStik 6F 5.5 fl oz ...... 27.50f-n Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 28.63f-n Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5 WDG 5.0 oz + Medallion 50WP 0.5 oz ...... 30.25f-n Insignia 20WDGG 0.7 oz + Iprodione Pro2SE 4.0 fl oz + Manicure Ultra 82.5WDG 5.0 oz ...... 31.50f-n Armada 50WP 1.2 oz FB Revere 4F 12.0 fl oz ...... 33.63f-n Tartan 288SC 2.0 fl oz + Chipco 26GT 6.0 fl oz ...... 33.75f-n Tartan 288SC 2.0 fl oz + Chipco 26GT 6.0 fl oz + SYNC ...... 34.63f-m DPX-LEM17 20SC 1.0 fl oz + DPX YT669 22.5SC 0.425 fl oz ...... 35.88f-l Spectator Ultra 1.3ME 4.0 fl oz + Insignia 20WDG 0.7 oz FB Manicure Ultra 82.5WDG 5.0 oz ... 36.50f-k Tartan 288SC 2.0 fl oz + Turfcide 400 6.0 fl oz ...... 41.25e-j Spectro 90WDG 4.0 oz + Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 41.50e-j Instrata 3.61SE 5.0 fl oz ...... 44.63e-i Spectro 90WDG 4.0 oz + Cleary 26/36 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz + SYNC ...... 45.38e-i Turfcide 400 12.0 fl oz ...... 46.25e-h Banner MAXX 1.3ME 2.0 fl oz + Turfcide 400 6.0 fl oz ...... 49.63d-g Spectator Ultra 1.3ME 4.0 fl oz FB Revere 4F 12.0 fl oz ...... 53.13c-f Banner MAXX 1.3ME 2.0 fl oz + Daconil WeatherStik 6F 5.5 fl oz ...... 53.75c-f DPX-H6573 37.7EC 0.44 fl oz ...... 65.00b-e DPX-LEM17 20SC 1.0 fl oz + DPX-H6573 37.7EC 0.44 fl oz ...... 66.25b-d DPX-LEM17 20SC 1.0 fl oz ...... 76.25a-d DPX-YT669 22.5SC 0.425 fl oz ...... 76.50a-d DPX-LEM17 20SC 0.5 fl oz ...... 79.00a-c Rapeseed 1 day germination 500g/100 ft2 ...... 81.25ab Mustard seed 1 day germination 100g/100 ft2 ...... 82.13ab Oilseed radish seed 1 day germination 500g/100 ft2 ...... 84.38ab Untreated Control ...... 91.75ab Rapeseed 1 day germination 100g/100 ft2 ...... 94.25a Rapeseed 6 day germination 500g/100 ft2 ...... 95.50a Oilseed radish seed 6 day germination 500 g/100 ft2 ...... 95.63a Mustard seed 6 day germination 500 g/100 ft2 ...... 96.25a Oilseed radish seed 1 day germination 100g/100 ft2 ...... 97.75a Mustard seed 1 day germination 500 g/100 ft2 ...... 98.75a_ LSD (α=0.05) ...... 27.42__

108

ANNUAL BLUEGRASS (Poa annua cv. unknown) T.D. Blunt and N. A. Tisserat Speckled snow mold (Typhula ishikariensis) Bioagricultural Sciences and Pest Management Gray Snow Mold (Typhula Incarnata) Colorado State University Fort Collins, CO 80523-1177

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2007-2008.

Experiments for control of speckled and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of several treatments of fungicides were applied 27 Sept 07. A second application of the initial treatments and the remainder of the treatments were applied 24 Oct 07. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 12 May 08.

Snow cover this field season was delayed. The first permanent snow was on or about 19 Nov 07. This season had greater snow cover than had been seen in the previous 3 years. The depth of the snow, at one time, was about 4 feet deep. Final snow melt from the trial plots was about 10 May 08, which was about two weeks later than normal. At evaluation time, there was still snow cover on shaded parts of the golf course. Due to the length and amount of snow cover, disease pressure was high. The untreated plots averaged 98% area damaged. Even though there were several fungicide combinations that provided adequate control (<10% damage) and there were a few combinations that were close to the 10% control, the majority of the fungicide combinations did not provide acceptable levels of control.

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Spectro 90WDG 4.0 oz FB Clearys 26/36 8.0 fl oz + CL-EXP-9 1.2 oz ...... 1.50i Spectro 90WDG 4.0 oz FB CL-EXP-9 1.2 oz + Endorse 2.5WP 4.0 oz ...... 2.25i Reserve 575SC 3.8 fl oz + Compass 50WDG 0.25 oz ...... 2.25i Spectro 90WDG 5.75 oz + CL-EXP-9 1.2 oz ...... 2.75i Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + CL-EXP-9 1.2 oz ...... 4.50hi Lynx 240SC 1.5 fl oz + Compass 50WG 0.25 oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 7.00g-i Instrata 3.61SE 9.3 fl oz + Medallion 50WP 0.25 oz ...... 7.00g-i Spectro 90WDG 4.0 oz FB Clearys 26/36 8.0 fl oz + Endorse 2.5WP 4.0 oz ...... 7.50g-i Reserve 575SC 7.6 fl oz + Chipco 26GT 4.0 fl oz ...... 10.50f-i Spectro 90WDG 4.0 FB Cleary 26/36 4.0 fl oz + Endorse 4.0 oz ...... 10.50f-i Instrata 3.61SE 11.0 fl oz + Medallion 50WP 0.25oz ...... 12.50f-i Insignia 20WDG 0.7 oz + Trinity 19.2SC 1.0 fl oz + Turfcide 400 6.0 fl oz ...... 13.75e-i Instrata 3.61SE 11.0 fl oz ...... 14.25e-i

109

Lynx 240SC 1.5 fl oz + Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 18.75d-i Insignia 20WDG 0.7 oz + Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 19.50d-i Trinity 19.2SC 1.0 fl oz + Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 20.00d-i Insignia 20WDG 0.7 oz + Chipco 26GT 4.0 fl oz + Turfcide 400 6.0 fl oz ...... 22.50d-i Banner MAXX 1.3ME 4.0 fl oz + Daconil Ultrex 82.5WDG 4.5 oz + Medallion 50WP 0.33 oz ...... 25.00c-h Instrata 3.61SE 5.5 fl oz FB Instrata 3.61 5.5 fl oz ...... 27.50c-g Instrata 3.61SE 7.0 fl oz FB Instrata 3.61SE 7.0 fl oz ...... 30.00c-f Instrata 3.61SE 9.3 fl oz ...... 31.25c-f Banner MAXX 1.3ME 3.25 fl oz + Daconil Ultrex 82.5WDG 4.5 oz + Medallion 50WP 0.27 oz .... 31.25c-f Tartan 288SC 2.0 fl oz + Turfcide 400 6.0 fl oz ...... 34.25c-e Insignia 20WDG 0.7 oz + Trinity 19.2SC 1.0 fl oz + Chipco 26GT 4.0 fl oz ...... 36.25cd Insignia 20WDG 0.7 oz + Trinity 19.2SC 1.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 37.25cd Tartan 288SC 2.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 45.75bc Reserve 575SC 7.6 fl oz ...... 59.50 Instrata 3.61SE 9.3 fl oz ...... 65.00b Untreated Control ...... 98.25a__ LSD (α=0.05) ...... 21.20__

110

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2008-2009.

Experiments for control of speckled and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of several treatments of fungicides were applied 26 Sept 08. A second application of the initial treatments and the remainder of the treatments were applied 29 Oct 08. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 27 April 09.

The first permanent snow was on or about 15 Nov 08. This season had as much snow cover as the previous year. At evaluation time, there was still snow cover on parts of the golf course. At the time of evaluation, there was still a small amount of snow on the trial plots that needed to be shoveled off in order to rate the effectiveness of the treatments. Disease pressure was high again this year. The untreated plots averaged 92% area damaged. Even though there were several fungicide combinations that provided adequate control (<10% damage) and there were a few combinations that were close to the 10% control, the majority of the fungicides or combinations did not provide acceptable levels of control.

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Spectro 90WDG 4.0 oz FB Cleary 26/36 8.0 fl oz + CL-EXP-9 1.2 oz ...... 4.50s Tartan 299SC 2.0 fl oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 5.25rs Spectro 90WDG 4.0 FB Instrata 3.61SE 11.0 fl oz + Medallion 50WP 0.25oz ...... 6.25rs Pegasus HPX 3.6 fl oz + Dovetail 39.3EC 4.0 fl oz + KestralMEX 1.3ME 3.0 fl oz + Disarm SC 0.36 fl oz ...... 6.50rs Triton Flo 367SC 0.85 fl oz + Compass 50WG 0.25 oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 6.50rs Reserve 4.8SC 5.4 fl oz + Compass 50WG 0.25 oz ...... 6.75q-s Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + CL-EXP-9 1.2 oz ...... 7.50q-s Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + Endorse 2.5WP 4.0 oz + CL-EXP-11 6.0 fl oz . 8.50p-s Instrata 3.61SE 8.0 fl oz + Headway 167EC 1.5 fl oz ...... 10.00o-s Headway 167EC 3.0 fl oz + Concert 4.3SC 8.25 fl oz ...... 12.00n-s USF26019T 6.0 fl oz + Triton Flo 367SC 0.85 fl oz ...... 12.00n-s Spectro 90WDG 4.0 oz FB CL-EXP-9 1.2 oz + Endorse 2.5WP 4.0 oz ...... 12.00n-s

111

Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + Endorse 4.0 2.5WP 4.0 oz ...... 12.25n-s Triton Flo 367SC 1.1 fl oz + Compass 50WG 0.2 oz + Daconil Ultrex 82.5WDG 5.0 oz ...... 12.75n-s Spectro 90WDG 5.75 oz + CL-EXP-9 1.2 oz ...... 13.00n-s USF26019T 5.0 fl oz + Triton Flo 367SC 0.85 fl oz ...... 13.25n-s Instrata 3.61SE 5.5 fl oz FB Instrata 3.61SE 5.5 fl oz ...... 15.00m-s Spectro 90WDG 4.0 oz FB Cleary 26/36 4.0 fl oz + CL-EXP-9 1.2 oz ...... 15.50m-s Curalan EG 50WP 1.0 oz + Daconil Ultrex 82.5WDG FB Insignia 20WDG 0.5 oz + Trinity 1.67SC 1.0 fl oz ...... 15.50m-s Banner MAXX 1.3ME 2.0 fl oz FB Instrata 3.61SE 11.0 fl oz + Medallion 50WP 0.25 oz ...... 18.00m-s Instrata 3.61SE 11.0 fl oz ...... 18.25m-s Banner MAXX 1.3ME 4.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz + Medallion 50WP + Chipco 26GT 4.0 fl oz ...... 18.75m-s Pegasus HPX 5.5 fl oz + Dovetail 39.3EC 4.0 fl oz + Disarm 0.25G 0.36 oz ...... 20.00l-s Banner MAXX 1.3 ME 4.0 fl oz + Medallion 50WP 0.4 oz + Chipco 26GT 4.0 fl oz ...... 22.50k-s USF26019T 4.0 fl oz + Triton Flo 367SC 0.85 fl oz ...... 27.25j-r Disarm 480SC 3.0 fl oz + Chipco 26GT 4.0 fl oz ...... 28.75j-r Instrata 3.61SE 9.3 fl oz ...... 30.00i-p Tourney 50WDG 0.44 oz + Cleary’s 3336 4.0 fl oz ...... 31.25i-o Curalan EG 50WG 1.0 oz + Daconil Ultrex 82.5WDG 3.2 oz FB Trinity 1.67SC 1.5 fl oz + Daconil Ultrex 82.5WDG 3.2 oz...... 33.75h-n Turfcide 400 6.0 fl oz + Spectro 90WDG 4.0 oz ...... 37.00g-m Tourney 50WDG 0.44 oz + Daconil Ultrex 82.5WDG 3.2 oz FB Tourney 50WDG 0.44 oz + Daconil Ultrex 82.5WDG 3.2 oz...... 42.00f-l Trinity 1.67SC 1.0 fl oz + Insignia 20WDG 0.5 oz ...... 42.00f-l AND3150 5 lb FB AND8076 5.95 lb + AND5017 6.66 lb ...... 42.50f-k Trinity 1.67SC 1.5 fl oz + Insignia 20WDG 0.5 oz ...... 48.00f-j Tourney 50WDG 0.44 oz FB Tourney 50WDG 0.44 oz ...... 51.25e-i AND5017 6.66 lb ...... 55.00e-h Turfcide 400 9.0 fl oz ...... 57.50d-g AND3150 5.0 lb FB AND5017 6.66 lb ...... 62.00c-f Turfcide 400 12.0 fl oz ...... 63.25c-f AND3150 3.75 lb FB AND5017 3.33 lb ...... 70.50b-e Disarm-G 4.5 lb ...... 72.75b-e AND5017 3.33 lb ...... 77.75a-d Disarm 480SC 0.36 fl oz...... 78.75a-d Insignia 20WDG 0.7 oz ...... 83.00a-c Rhapsody 60EC 5.0 fl oz + Insignia 20WDG 0.35 oz ...... 84.00a-c Turfcide 400 6.0 fl oz ...... 86.75ab Untreated Control ...... 92.25ab Untreated Control ...... 96.00a__ LSD (α=0.05) ...... 22.25___

112

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2009-2010.

Experiments for control of speckled and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 14, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of several treatments of fungicides were applied 2 Oct 09. A second application of the initial treatments and the remainder of the treatments were applied 05 Nov 09. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 10 May 10.

Snow cover this field season was delayed. The first permanent snow was on or about 19 Nov 10. First snow melt was in Apr 10 with additional snow cover after the first melt. Final snow melt from the trial plots was about 06 May 10, which was about two weeks later than normal. At evaluation time, there was still snow cover on shaded parts of the golf course. Disease pressure on the fairway was consistent with previous years. Even though there were several fungicide combinations that provided adequate control (<20% damage) and there were a couple combinations that were close to the 10% control threshold, about half of the treatment combinations did not provide acceptable levels of control.

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Reserve 4.8SC 5.4fl oz + Compass 50WG 0.25 oz ...... 9.67h Instrata 3.61SE 5.5 fl oz FB Instrata 3.61SE 5.5 fl oz ...... 10.00h Instrata 3.61SE 5.5 fl oz + Fore 80WP 8.0 oz FB Instrata 3.61SE 5.5 fl oz + Fore 80WP 8.0 oz ..... 10.33gh Insignia 20WDG 0.54 oz + Trinity 1.67SC 1.0 oz + Iprodione Pro 2SE 4.0 fl oz ...... 11.00gh Tourney 50WDG 0.37 oz + Cleary 3336 4.0 fl oz FB Tourney 50WDG 0.37 oz + Cleary 3336 4.0 fl oz ...... 11.67gh Instrata 3.61SE 11.0 fl oz ...... 11.67gh Interface 24.5SC 4.0 fl oz + Turfcide 400 8.0 fl oz ...... 12.67f-h Interface 24.5SC 5.0 fl oz + Triton Flo 367SC 0.85 fl oz...... 12.67f-h Daconil Ultrex 82.5WDG 5.5 oz + Medallion 50WP 0.50 oz + Banner MAXX 1.3ME 4.0 fl oz ...... 12.67f-h Interface 24.5SC 6.0 fl oz + Triton Flo 367SC 0.85 fl oz...... 14.33f-h V10277 45G 2.4 lb + Cleary 3336 4.0 fl oz FB V10277 45G 2.4 lb + Cleary 3336 4.0 fl oz ...... 15.00f-h Turfcide 400 16.0 fl oz ...... 16.00e-h Insignia 20SC 0.54 oz + Trinity 1.67SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 16.67e-h

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Reserve 4.8SC 4.5 fl oz + Compass 50WG 0.20 oz ...... 16.67e-h Kestrel MEX 1.3ME 4.0 fl oz + Pegasus HPX 5.5 fl oz FB Kestrel MEX 1.3ME 4.0 oz + Raven 2SC 4.0 fl oz ...... 16.67e-h Daconil Ultrex 82.5WDG 3.2 oz + Medallion 50WP 0.50 oz + Banner MAXX 1.3ME 4.0 fl oz ...... 16.67e-h Turfcide 400 12 fl oz + Rhapsody 60EC 5.0 fl oz ...... 18.33e-h Peregrine 66WDG 4.0 oz + Pegasus HPX 3.0 fl oz + Kestrel MEX 1.3ME 4.0 fl oz + Raven 2SC 4.0 fl oz ...... 18.33e-h Instrata 3.61SE 11.0 fl oz + Medallion 50WP 0.17 oz ...... 19.33e-h Interface 24.5SC 4.0 fl oz + Triton Flo 367SC 0.85 fl oz...... 20.00e-h Honor 28WG 0.83 oz + Trinity 1.67SC 1.0 oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 20.00e-h Insignia 20WG 0.7 oz + Tourney 50WDG 0.37 oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 21.67e-h Curalan EG 50WDG 1.0 oz + Daconil Ultrex 82.5WDG 3.2 oz FB Insignia 20WG 0.7 oz + Trinity 1.67SC 1.0 oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 23.33d-h Reserve 4.8SC 4.5 fl oz + Compass 50WG 0.25 oz ...... 23.33d-h Disarm 480SC 5.8 fl oz + Raven2SC 4.0 fl oz ...... 23.33d-h V10277 45G 2.4 lb FB V10277 45G 2.4 lb ...... 26.67d-h Chipco 26GT 4.0 fl oz + Medallion 50WP 0.5 oz + Banner MAXX 1.3ME 4.0 fl oz ...... 26.67d-h Kestrel MEX 1.3ME 4.0 fl oz FB Disarm 480SC 5.8 fl oz + Raven 2SC 4.0 fl oz ...... 28.33d-g Chipco 26GT 4.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz + Medallion 50WP 0.5 oz + Banner MAXX 1.3ME 4.0 fl oz ...... 28.33d-g Disarm M 1.0 fl oz + Pegasus DF 3.6 fl oz ...... 30.00d-f Daconil Ultrex 82.5WDG 5.5 oz + Medallion 50WP 0.25oz + Banner MAXX 1.3ME 4.0 fl oz ...... 34.00de Pegasus L 3.6 fl oz + Kestrel MEX 1.3ME 3.0 fl oz + Dovetail 39.3EC 4.0 fl oz + Disarm 480SC 0.36 fl oz...... 40.00cd Tourney 50WDG 0.37 oz FB Tourney 50WDG 0.37 oz ...... 53.33bc Rhapsody 60EC 5.0 fl oz ...... 58.33b Untreated Control ...... 81.67a__ LSD (α=0.05) ...... 18.16___

114

Fungicide evaluation for the preventive control of snow molds on Annual Bluegrass, 2010-2011.

Experiments for control of speckled and gray snow mold were performed on an Annual Bluegrass fairway at Vail Golf Club, Fairway 16, Vail, CO (average elevation 8150 ft.). Treatments were arranged in a randomized complete block design with four replications. Individual plots measured 5 ft. X 5 ft. First applications of several treatments of fungicides were applied 1 Oct 10. A second application of the initial treatments and the remainder of the treatments were applied 02 Nov 10. All fungicides were applied in the equivalent of 5 gal. water/1000 sq. ft. A CO2-powered sprayer equipped with 8003 Tee-jet nozzles delivered the treatments at 30 psi. Fungicide effectiveness was evaluated after snowmelt on 06 May 10.

Snow cover this field season was earlier than in previous years. The first snow was on or about 25 Oct 10. Final snow melt from the trial plots was about 06 May 11, which was about two weeks later than normal. At evaluation time, there was still snow cover on shaded parts of the golf course. This year we had a La Niña weather pattern which provided a total of 524 inches of snow over the entire season. Disease pressure on the fairway was consistent with previous years. The majority of the treatments provided control of <20% disease incidence.

Table 1. Snow mold fungicide trials at Vail Golf Club, Vail, CO______Treatment and Rate/1000 sq. ft. (unless otherwise noted) Disease Rating_ Interface 24.5SC 6.0 fl oz + Triton Flo3667SC 0.85 fl oz ...... 0.00g Pegasus HPX 2.75 fl oz + Kestrel MEX 1.3ME 2.0 fl oz + Dovetail 39.3 4.0 fl oz + Disarm 480SC 0.18 fl oz FB Pegasus HPX 2.75 fl oz + Kestrel MEX 1.3ME 2.0 fl oz + Dovetail 39.3 4.0 fl oz + Disarm 480SC 0.18 fl oz ...... 0.00g Torque 3.6SE 0.9 fl oz + Clearys 26/36 4.0 fl oz ...... 0.75g Instrata 3.61SE 5.5 fl oz FB Instrata 3.61SE 5.5 fl oz ...... 0.75g Reserve 4.8SC 5.4 fl oz + Interface 24.5SC 6.0 fl oz ...... 1.00g Torque 3.6SE 0.6 fl oz + Clearys 26/36 4.0 fl oz ...... 1.00g Instrata 3.61SE 5.0 fl oz + Medallion 50WP 0.25 oz + Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5WDG 2.0 oz...... 1.25g Peregrine 66WDG 6.0 oz + Kestrel MEX 1.3ME 2.0 fl oz FB Peregrine 66WDG 6.0 oz + Kestrel MEX 1.3ME 2.0 fl oz ...... 1.25g Instrata 3.61SE 11.0 fl oz + Medallion 50WP 0.25 oz ...... 1.75g Instrata 3.61SE 11.0 fl oz ...... 3.00e-g Torque 3.6SE 0.6 fl oz + Affirm WDG 0.9 oz + Spectro 90WDG 3.67 oz ...... 3.75e-g

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Instrata 3.61SE 9.0 fl oz + Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5WDG 2.0 oz ...... 3.75e-g Pegasus HPX 5.50 fl oz + Kestrel MEX 1.3ME 4.0 fl oz + Dovetail 39.3 8.0 fl oz + Disarm 480SC 0.36 fl oz...... 4.00e-g Reserve 4.8SC 5.4 fl oz + Compass 50WG 0.25 oz ...... 4.50e-g Torque 3.6SE 0.6 fl oz + Affirm WDG 0.9 oz ...... 4.50e-g Instrata 3.61SE 9.0 fl oz + Medallion 50WP 0.25 oz + Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5WDG 2.0 oz...... 5.00e-g Curalan EG 50WG 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz FB Insignia 20WDG 0.54 oz + Trinity 1.67SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 5.25e-g Torque 3.6SE 0.6 fl oz + Clearys 26/36 4.0 fl oz + Spectro 90WDG 3.67 oz ...... 6.25e-g Interface 24.5SC 3.0 fl oz + Triton Flo 367SC 0.5 fl oz FB Interface 24.5SC 3.0 fl oz + Triton Flo 367SC 0.5 fl oz ...... 6.25e-g Torque 3.6SE 0.9 oz + Clearys 26/36 4.0 fl oz + Spectro 90WDG 3.67 oz ...... 7.50e-g Insignia 20WDG 0.7 oz + Trinity 1.67SC 1.5 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 8.50e-g Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5WDG 5.5 oz...... 8.75e-g Interface 24.5SC 5.0 fl oz + Reserve 4.8SC 5.4 fl oz ...... 11.25e-g Spectro 90WDG 5.0 oz ...... 19.25d-f Insignia 20WDG 0.54 oz + Trinity 1.67SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 20.00c-e Instrata 3.61SE 5.0 fl oz + Banner MAXX 1.3ME 2.0 fl oz + Daconil Ultrex 82.5WDG 2.0 oz ...... 32.50cd Honor 28WG 0.84 oz + Trinity 1.67SC 1.0 fl oz + Daconil Ultrex 82.5WDG 3.2 oz ...... 34.50cd Torque 3.6SE 0.9 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 36.25c Torque 3.6SE 0.6 fl oz + Daconil Ultrex 82.5WDG 5.5 oz ...... 60.00b Untreated Control ...... 90.00a__ LSD (α=0.05) ...... 16.37___

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APPENDIX 3

OPTIMIZING THE GC/MS PROTOCOL

117

The process of optimizing the GC/MS protocol using chlorothalonil analytical standards was started in August of 2005. The objective was to determine the amount of chlorothalonil recovery we could expect. Initially, the concentrations used were 10, 25,

50, 75 and 100 ng/ml dilutions of a 0.1 mg/ml analytic standard of chlorothalonil in acetylnitrile combined with 10 µl of a 0.5 ng/ml butylate internal standard stock solution

(50 µl of butylate in 100 ml toluene). This initial run on the GC/MS was satisfactory and the process continued. The analytical standards along with the internal standards were run three separate times and worked well, so the next step was to see which solvent(s) would work best to extract chlorothalonil from leaf tissue. Since chlorothalonil is a non- polar solution, water is the extraction medium of choice to get it off the turf tissue, however, water cannot be used in the GC/MS runs. Toluene is generally the solvent of choice for the machine. A mixture of acetone or methanol and water at various percentages (25, 50 or 75%) as first solvent solutions were attempted. After the extraction process, the acetone or methanol was evaporated using a RapidVac at 50°C,

60 vortex speed, then adding toluene to the remaining water to extract the chlorothalonil into the toluene. These results were acceptable, but not good. The next step was to try acidifying the solution of acetone and water using 0.5% phosphoric acid. With this extraction method, we only recovered approximately 80% of the chlorothalonil standard.

Using acetone as an extraction solvent also increased the amount of chlorophyll extracted when using inoculated turf. After experimenting with several iterations of acetone/water or methanol/water extraction methods, it was determined that neither acetone or methanol would be an acceptable solvent to extract chlorothalonil. Also, because of the increased amount of chlorophyll extracted along with the chlorothalonil,

118 the results of acetone and methanol were not acceptable. Also, in the presence of acetone and acetonitrile (MeCN), chlorothalonil would degrade in the clear sampling vials but remain stable in amber colored vials (Mastovska and Lehotay, 2004). After experimenting with several iterations of acetone/water and methanol/water using inoculated turf and tomato plants, and still having to use toluene as the end solvent for the GC runs, it was decided to try extracting directly into toluene. Since the vials used for sampling were clear, toluene was the superior exchange solvent. It was also determined that the starting weight of 1g of turf tissue was too large a sample size. In optimizing the protocol, it was determined that 0.20g turf tissue in 10 ml diH2O and 10 ml toluene would extract an amount of chlorothalonil that would not overwhelm the

GC/MS and would produce consistent results.

119